RapidFire 400 System User Guide
Agilent, RapidFire, 400, System, User, Guide
Agilent;, RapidFire;, 400;, System;, User;, Guide
Agilent Technologies
RapidFire 400 System User Guide
D0004197 RapidFire400UserGuideA02 RapidFire 400 System
User Guide
Original instructions
Notices
Document Identification
D0004197 July 2021 Revision A.02
Copyright
© Agilent Technologies, Inc. 2021
No part of this manual may be reproduced in any form or by any means (including electronic storage and retrieval or translation into a foreign language) without prior agreement and written consent from Agilent Technologies, Inc. as governed by United States and international copyright laws.
Agilent Technologies, Inc. 5301 Stevens Creek Blvd. Santa Clara, CA 95051
Software Revision
This guide is valid for the C.01.00 revision or higher of the PRODUCT NAME program and compatible PRODUCT NAME programs, until superseded.
Instrument Manufacturing
Manufactured by Agilent Technologies Singapore Pte. Ltd. No. 1 Yishun Avenue 7, Singapore 768923
Temperature Limits
Operating Temperature: 15°C to 35°C Storage Temperature: -40°C to 70°C
Software Manufacturing
Manufactured for Agilent Technologies 5301 Stevens Creek Blvd Santa Clara, CA 95051
Warranty
The material contained in this document is provided "as is," and is subject to being changed, without notice, in future editions. Further, to the maximum extent permitted by applicable law, Agilent disclaims all warranties, either express or implied, with regard to this manual and any information contained herein, including but not limited to the implied warranties of merchantability and fitness for a particular purpose. Agilent shall not be liable for errors or for incidental or consequential damages in connection with the furnishing, use, or performance of this document or of any information contained herein. Should Agilent and the user have a separate written agreement with warranty terms covering the material in this document that conflict with these terms, the warranty terms in the separate agreement shall control.
Technology Licenses
The hardware and/or software described in this document are furnished under a license and may be used or copied only in accordance with the terms of such license.
Restricted Rights Legend
U.S. Government Restricted Rights. Software and technical data rights granted to the federal government include only those rights customarily provided to end user customers. Agilent provides this customary commercial license in Software and technical data pursuant to FAR 12.211 (Technical Data) and 12.212 (Computer Software) and, for the Department of Defense, DFARS 252.227-7015 (Technical Data Commercial Items) and DFARS 227.7202-3 (Rights in Commercial Computer Software or Computer Software Documentation).
Safety Notices
CAUTION
A CAUTION notice denotes a hazard. It calls attention to an operating procedure, practice, or the like that, if not correctly performed or adhered to, could result in damage to the product or loss of important data. Do not proceed beyond a CAUTION notice until the indicated conditions are fully understood and met.
WARNING
A WARNING notice denotes a hazard. It calls attention to an operating procedure, practice, or the like that, if not correctly performed or adhered to, could result in personal injury or death. Do not proceed beyond a WARNING notice until the indicated conditions are fully understood and met.
Content
1 Before you start 1
About this guide 2 Related guides and more information 3 Safety notes 3 Making an emergency stop 4 About the RapidFire 400 System 5 Hardware overview 6 Materials supplied 8
2 System preparation 11
Preparing solvents 12 Checking waste capacity 12 Starting the system 13 Loading and homing the cartridge changer 16 Homing the stage 18 Setting the stage teachpoints 18 Setting up the system for teaching and teaching the four touchpoints 20 Opening the Sipper Configuration Wizard and installing the teaching jig 21 Setting the Sipper Safe Height and teaching the four touchpoints 24 Preparing to verify the calculated teachpoints 30 Accepting or editing the calculated teachpoints 30 Verifying the calculated teachpoints 34 Checking the nanovalves 45 Preparing the peristaltic pump 47 Preparing the quaternary pumps 51 Preparing the sipper 52 Preparing the temperature control unit 53 Preparing the TOF or Q-TOF mass spectrometer 53 Preparing the TQ or Ultivo TQ mass spectrometers 61 Minimizing the required sample volume 62 Shutting down the RapidFire system between runs 64
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Content
3 Operating the RapidFire 400 in Plates mode 65
About Plates mode 66 Workflow for using Plates Mode 66 Before you start 66 Recommendations for preparing the plates 67 Loading a single plate 68 Loading multiple plates using the plate handler 68 Preparing the TOF or Q-TOF mass spectrometer 70 Preparing the TQ or Ultivo TQ mass spectrometer 72 Setting up the RapidFire parameters for a Plates mode run 74 Starting a run in Plates mode 78 Pausing a run in Plates mode 79 Stopping a run in Plates mode 79 Maintaining a log book of data 80
4 Operating the RapidFire 400 in Sequences mode 81
About Sequences mode 82 Workflow for operating in Sequences mode 83 Before you start 84 Creating a RapidFire Method file (.rfcfg) 84 Creating a RapidFire Sequence PlateMap file (.rfcfg) 88 Creating a RapidFire Batch file 90 Recommendations for preparing the sample plates 95 Loading a single plate 95 Loading multiple plates 96 Preparing the TOF or Q-TOF mass spectrometer 97 Preparing the TQ or Ultivo TQ mass spectrometer 98 Starting a run in Sequences mode 98 Using an information management file for a Sequences mode run 99 Creating a map file using MapFile Generator utility 100 Starting the run in Sequences mode using an information management file 108 Pausing a run in Sequences mode 110 Stopping a run in Sequences mode 111
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Maintaining a log book of data 111
5 Maintenance 113 Daily maintenance 114 Weekly maintenance 114 Shutting down the RapidFire system completely 114
6 RapidFire System flow path reference 117 State 1: Aspirate flow path 118 State 2: Wash/Load flow path 119 State 3: Extra Wash flow path 120 State 4: Elute flow path 121 State 5: Re-equilibrate flow path 122 Flush the sipper flow path 123 Physical colors of the tubing 124 Valve positions 124
A Changing the sequence of the sipper operation 125 About the sip sequence operation 126 Changing the sequence of sip operation 127
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Before you start
This section provides an overview of the RapidFire 400 System, where to find more information, safety notes, and materials supplied. It contains the following topics:
About this guide 2 Related guides and more information 3 Safety notes 3
Making an emergency stop 4 About the RapidFire 400 System 5 Hardware overview 6
Materials supplied 8
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Before you start
About this guide
About this guide
Who should read this guide
This guide is for people with the following job roles:
Table 1. Job roles and responsibilities
Job role Installer Integrator Lab manager, administrator, or technician
Operator
Responsibilities
Unpacks, installs, and tests the device before it is used.
Configures hardware and writes software.
· Manages the automation system that contains the device. · Develops the applications that are run on the system. · Develops training materials and standard operating procedures for
operators.
Performs the daily production work on the device and solves routine problems.
What this guide covers
This guide covers the description, setup, operation, and maintenance of the Agilent G9532A and G9532AA RapidFire 400 System.
This guide does not provide instructions for unpacking, installation, and data analysis.
What is new in this guide
Table 2. What is new in this guide
Change
Described in...
Release of MassHunter 11.0 (MH11) for TOF/QTOF. MH11 uses components of the OpenLab software which affect certain procedures and the user interface.
· "Preparing the TOF or Q-TOF mass spectrometer" on page 53. · "Preparing the TOF or Q-TOF mass spectrometer" on page 70. · "Preparing the TOF or Q-TOF mass spectrometer" on page 97
New default sequence of sip operation
"Changing the sequence of the sipper operation" on page 125
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Before you start
Related guides and more information
Software version and compatibility
This guide documents RapidFire Acquisition software v 6.0.
RapidFire Acquisition software 6.0 requires the Microsoft Windows 10 64-bit operating system.
RapidFire Acquisition software 6.0 is compatible with: · MassHunter Acquisition software 10.1 for TOF, Q-TOF, and TQ · MassHunter Acquisition software 1.2 for Ultivo
Related guides and more information
Use this guide in conjunction with the following: · RapidFire 400 System Safety Guide. Describes the potential safety hazards
and how to avoid them. · RapidFire System Data Analysis Guide. Describes how to process data
acquired from the RapidFire System using RapidFire Integrator or MassHunter Quantitative Analysis. · Agilent web site: www.agilent.com.
Safety notes
WARNING
Using controls, making adjustments, or performing procedures other than those specified in the user documentation can expose you to moving-parts hazards and hazardous voltage. Before using the RapidFire 400 System, make sure you are aware of the potential hazards and understand how to avoid being exposed to them.
Ensure you have read the RapidFire 400 System Safety Guide and are trained in the safe operation of the device.
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Making an emergency stop
Making an emergency stop
CAUTION
Use the emergency stop only for emergencies. If you stop a run with the emergency stop, you cannot continue the run. Otherwise, use one of the following procedures to stop the RapidFire System.
"Pausing a run in Plates mode" on page 79 "Stopping a run in Plates mode" on page 79
Procedure
To stop in an emergency: 1 Press the red emergency stop button on the front of the RapidFire system.
Power is cut to the motors and there is no further motion.
2 To restart the system:
a Release the emergency stop by twisting the knob while applying light pressure.
b Restart the run to continue.
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About the RapidFire 400 System
About the RapidFire 400 System
Product description
The RapidFire 400 System is a sample purification and injection system that interfaces directly with the mass spectrometer. The RapidFire System can simultaneously analyze multiple analytes in complex mixtures and remove many contaminants contained in those mixtures.
An optional temperature control unit enables the system to keep plates chilled while stored.
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Before you start
Hardware overview
Hardware overview
Primary hardware components
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Hardware overview
Table 3. RapidFire 400 System components
Item Name
Description
1 RapidFire computer and
monitor
Runs the RapidFire software and mass spectrometer software and communicates to the hardware. The monitor displays the user interface of the controlling and acquiring software.
2 Controller
Controls the chiller, if present, robot, and barcode reader stage.
3 Controller
Controls the plate stage, cartridge changer, and valves.
4 Ultivo Triple Quadrupole
Analyzes RapidFire samples.
LC/MS (not included with the
RapidFire 400 System)
5 Peristaltic pump
Pumps solvent to sipper wash station.
6 Quaternary pumps
Pump mobile phase throughout the system.
7
Pump with Quiet Cover for
Provides vacuum for the mass spectrometer.
Ultivo Triple Quadrupole
LC/MS (not included with the
RapidFire 400 System)
8 Wheels
Locking wheels to enable system mobility. To lock wheels, step down on the locking tab. To unlock, lift up the locking tab.
9 Solvent and waste secondary Holds the waste and liquids that supply the system.
container
10 Emergency stop button
Stops the motion of the system in an emergency.
11 Doors with safety interlock Protects you from the moving parts of the system.
12 RapidFire 400 frame
Houses the RapidFire 400 System.
13 Temperature control unit
(optional)
Maintains the temperature of sample plates (4° C to ambient)
14 Stacker
Holds the sample plates and moves them into robot-accessible position.
15 Plate handler
Moves the sample plates between the stacks, the barcode stage, and the plate stage.
16 Nanovalves
Direct the flow of mobile phase and samples throughout the system.
17 Plate stage
Controls x, and y-axis movement of plates to facilitate loading and unloading of plates and sipper access to wells.
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Materials supplied
Table 3. RapidFire 400 System components
Item Name
18 Barcode scanner stage 19 Barcode scanner
Description Holds the sample plate in position to enable scanning. Scans the sample plate.
CAUTION
All interconnecting cables in the RapidFire 400 System must be less than three meters.
Materials supplied
Agilent-supplied items
RapidFire System contents: · RapidFire 400 System instrument · Optional. Temperature control unit · Three digitally controlled high-pressure fluidic pumps · Digital peristaltic pump · Two barcode scanners · Waste collection vessels · 12-slot cartridge changer · Sip sensor Consumables: · Standard RapidFire solid-phase extraction (SPE) cartridges (see the table below)
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Materials supplied
WARNING
Customer-supplied items
· Assay-specific solvents, LC/MS grade, filtered by the manufacturer
Improper handling of solvents can result in personal injury. Handle solvents safely as described in the Agilent G9532A RapidFire 400 Safety Guide. Contact the manufacturer for safety data sheets (MSDS) for all materials that you use.
· 1-liter graduated cylinder · Pipettes and pipette tips · Sample 96-, 384- and/or 1536-well plates (with or without standard bar
codes)
RapidFire system cartridges
There are a number of different types of RapidFire cartridges available from Agilent, some of which are supplied with the instrument. See the RapidFire System ordering catalog for details.
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System preparation
This section describes how to prepare the RapidFire 400 System for a run. It contains the following topics: Preparing solvents 12 Checking waste capacity 12 Starting the system 13 Loading and homing the cartridge changer 16 Homing the stage 18 Setting the stage teachpoints 18 Setting up the system for teaching and teaching the four touchpoints 20 Opening the Sipper Configuration Wizard and installing the teaching jig 21 Setting the Sipper Safe Height and teaching the four touchpoints 24 Preparing to verify the calculated teachpoints 30 Accepting or editing the calculated teachpoints 30 Verifying the calculated teachpoints 34 Checking the nanovalves 45 Preparing the peristaltic pump 47 Preparing the quaternary pumps 51 Preparing the sipper 52 Preparing the temperature control unit 53 Preparing the TOF or Q-TOF mass spectrometer 53 Preparing the TQ or Ultivo TQ mass spectrometers 61 Minimizing the required sample volume 62 Shutting down the RapidFire system between runs 64
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System preparation
Preparing solvents
Preparing solvents
WARNING
Use only LCMS-grade, filtered, degassed solvents. Change solvents according to your assay.
Improper handling of solvents can result in personal injury. Handle solvents safely as described in the RapidFire 400 System Safety Guide. Contact the manufacturer for safety data sheets (MSDS) for all materials that you use.
To prepare the solvents: 1 Check that you have enough solvent for the runs you plan to make and fill the
following containers accordingly: · Lines 1A, 1B, 1C, 1D feed Pump 1 · Lines 2A, 2B, 2C, 2D feed Pump 2 · Lines 3A, 3B, 3C, 3D feed Pump 3 · Water in a bottle labeled 100% water · Acetonitrile in a bottle labeled 100% acetonitrile · Other solvents such as methanol, in a labeled bottle 2 Gently shake the filters and whirl the flask around to remove air bubbles.
Checking waste capacity
WARNING
Discard all waste according to your lab's waste disposal procedures and in compliance with all local, state, and national safety regulations.
To check waste capacity:
1 Check that the waste containers have enough room to accommodate the waste that will be generated from the runs you have planned.
2 If more capacity is required:
a Empty the waste carboy where the two green system waste lines are directed.
b Empty the vacuum waste flask.
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Starting the system
c Check that the two green waste lines are not dipping in waste fluid, but merely sticking out of the carboy lid.
Starting the system
To start the system:
1 Press the two green power buttons on the back of the frame to turn on the system power.
2 Turn on the quaternary pumps, peristaltic pump, temperature control unit (if present), computer, and vacuum source.
3 Double-click the RapidFire Control Panel
desktop icon.
4 Click File > Choose CFGs.
5 Select the configuration folder under C:\Agilent\RapidFire.
6 Click Start in the RapidFire Control Panel.
7 Double-click the RapidFire UI
desktop icon.
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Starting the system
8 Click System Tools > Network Settings.
9 Verify that the IP addresses for the RapidFire computer and the mass spectrometer computer are correct:
NOTE
To find the computer IP address: a Click the Start icon in the Windows desktop, type Run, and select the
Command Prompt App. b In the Command Prompt window, type ipconfig-all and press Enter.
If you are controlling the mass spectrometer and RapidFire on one computer, their IP addresses will be the same.
10 Click Connect in the main window to connect to the program server. The indicator lights next to Connect turn green.
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Starting the system
If one indicator is red, the system is connected but the door is open or the emergency stop button is pressed.
If both lights are red, the system is not connected.
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System preparation
Loading and homing the cartridge changer
Loading and homing the cartridge changer
To load and home the cartridge changer:
1 Click System Tools > Cartridge Changer to open the Cartridge Changer dialog box.
2 Select Load and click Go To to move the changer to the load position.
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NOTE
System preparation Loading and homing the cartridge changer
3 Load up to 12 new cartridges in the holder:
Always put one cartridge in slot 1 as the cartridge changer travels to this position after the homing routine is complete.
a Remove the protective packaging and insert the new cartridges in the slots, with the notch end facing up.
Figure 1. Cartridge changer with 12 cartridges
b Update the information in the Cartridge Changer dialog box. c To reset the Sample Count of a particular cartridge to zero or any given
non-zero value, double-click the corresponding field and confirm your input. d Click Update. e In the Cartridge Changer dialog box, select Home from the Go To list and click Go To to home the changer.
The Homed indicator turns green.
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Homing the stage
Homing the stage
After you set the stage teachpoints for your particular plate type, you only need to home the stage when performing a run.
To home the stage:
1 Click System Tools > Sipper Configuration to open the Sipper Configuration Wizard.
2 Make sure the plate type you are using is selected in the plate to configure field.
3 Click Home and wait for all three axis stages (x, y, and z) to get to home position (0,0,0).
4 Do one of the following: · If you have not yet set the stage teachpoints, go to "Setting the stage teachpoints" on page 18. · If you have set the stage teachpoints, close the wizard and go to "Checking the nanovalves" on page 45 to continue with preparing for a run.
Setting the stage teachpoints
About setting the stage teachpoints
You teach the RapidFire system stage positions when you do the following: · Use a new plate type · Replace the sipper tube or guiding needle
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NOTE
System preparation Setting the stage teachpoints
Setting the stage teachpoints for a plate requires that the plate is first predefined in the software. The RapidFire software has four predefined labware types (.platecfg files) that can be used to perform the teaching: · 96-well standard microplate · 96-deep well microplate · 384-well standard microplate · 1536-well standard microplate
Using 1536-well microplates requires smaller diameter injection loop and sipper tube configuration to minimize sample volume. See "Minimizing the required sample volume" on page 62 for more information.
About the Sipper Configuration Wizard
The software uses the data from the first four steps of the wizard (Touchpoint 1, Touchpoint 2, Touchpoint 3, and Touchpoint Z) to calculate all of the stage teachpoints including the plate teachpoints, wash station teachpoints, and matrix teachpoints.
Once you click Set for the calculated teachpoints, you can verify and adjust the calculated teachpoints by continuing with the wizard, or you can skip the verification by clicking Next, without clicking Set, in the relevant dialog box.
You also have the option of setting all the teachpoints manually by skipping the first four steps of the wizard. To skip a step, click Next in the dialog box.
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Setting up the system for teaching and teaching the four touchpoints
Before you begin
Make sure you have the following: · An empty plate of the type you are teaching and, if applicable, four empty
matrix bottles · Stage teaching jig and attachment wire with clip
Use the following workflow to set the stage teachpoints.
Step Task
See
1
Set up the teaching jig and teach the four
touchpoints.
"Setting up the system for teaching and teaching the four touchpoints" on page 20
2
View and accept or edit the calculated
teachpoints.
"Accepting or editing the calculated teachpoints" on page 30
3
Verify the wash station, corner well, and matrix "Verifying the calculated teachpoints" on
position teachpoints.
page 34
Setting up the system for teaching and teaching the four touchpoints
In this first part of the wizard, you will do the following:
Step Task
1
Open the wizard and install the teaching jig.
2
Set the Sipper Safe Height and teach the four
touchpoints.
3
Prepare to verify the calculated teachpoints.
See
"Opening the Sipper Configuration Wizard and installing the teaching jig" on page 21
"Setting the Sipper Safe Height and teaching the four touchpoints" on page 24
"Preparing to verify the calculated teachpoints" on page 30
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Opening the Sipper Configuration Wizard and installing the teaching jig
Opening the Sipper Configuration Wizard and installing the teaching jig
To open the wizard and install the jig:
1 Open the Sipper Configuration Wizard (see "Homing the stage" on page 18) and home the stage.
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Opening the Sipper Configuration Wizard and installing the teaching jig
2 Install the stage teaching jig:
a Open the doors to disengage the stage motors.
b Place the teaching jig on the stage and make sure teachpoints labeled 1 and 3 on the jig are near the A1 corner of the stage.
c Connect one end of the jig wire with the connector pin to the jig itself.
d With the alligator clip, attach the other end of the jig wire to the sipper assembly near the top of the needle.
e Turn on the power to the jig.
3 If you are using matrix bottles, place them in the four holes marked M.
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Opening the Sipper Configuration Wizard and installing the teaching jig
4 Select the plate type from the plate to configure list to activate the wizard.
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Setting the Sipper Safe Height and teaching the four touchpoints
Setting the Sipper Safe Height and teaching the four touchpoints
CAUTION
Set the Sipper Safe Height before starting operation to avoid damage to the equipment.
NOTE
To set the Sipper Safe Height and teach the four touchpoints: 1 Set the Sipper Safe Height as follows:
a Turn the black knob on the end of the z-stage to adjust the z-axis so that the bottom of the sipper clears all potential obstacles (such as the plate, wash station, and matrix bottles) by about 0.5 cm (1/4 inch).
Perform this step regardless of where the stage moves along the x- or y-axis.
b Close the door to engage the stage motors. c Click Set and then Next in the Sipper Configuration Wizard.
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Setting the Sipper Safe Height and teaching the four touchpoints
2 Configure Touchpoint 1 Position:
a Open the doors and manually adjust the position of the stage in the x-, yand z-axes so that the sipper is adjacent to the Touchpoint 1 mark on the jig, as the following figure shows.
b Continue to adjust the x-axis (side-to-side) of the stage until the sipper tip contacts the teaching jig and causes the light to flicker and buzzer to sound intermittently on the jig.
Figure 2. Location of Touchpoint 1 position
c Close the door to engage the stage motors. d Click Set and then Next in the Sipper Configuration Wizard.
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Setting the Sipper Safe Height and teaching the four touchpoints
3 Configure Touchpoint 2 Position:
a Open the doors to disengage the motors.
b Manually adjust the x-, y-, and z-axes of the stage so that the sipper is adjacent to the Touchpoint 2 mark on the jig, as the following figure shows.
c Continue to adjust the x-axis of the stage until the sipper tip contacts the teaching jig and causes the light to flicker and buzzer to sound intermittently on the jig.
Figure 3. Location of Touchpoint 2 position
d Close the door to engage the stage motors. e Click Set and then Next in the Sipper Configuration Wizard.
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Setting the Sipper Safe Height and teaching the four touchpoints
4 Configure Touchpoint 3 Position:
a Open the doors to disengage the motors.
b Manually adjust the x-, y-, and z-axes of the stage so that the sipper is adjacent to the Touchpoint 3 mark on the jig, as the following figure shows.
c Continue to adjust the y-axis backward and forward of the stage until the sipper tip contacts the teaching jig and causes the light to flicker and buzzer to sound intermittently on the jig.
Figure 4. Location of Touchpoint 3 position
d Close the door to engage the stage motors. e Click Set and Next in the Sipper Configuration Wizard.
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Setting the Sipper Safe Height and teaching the four touchpoints
5 Configure the Touchpoint Z Position:
a Open the doors to disengage the motors.
b Remove the jig from the stage.
c Manually adjust the x-, y-, and z-axes so that the sipper is above the empty stage.
d Slowly lower the sipper in the z-axis (vertical) until the tubing extending over the tip of the sipper needle touches the surface of the stage (1) and the tubing flexes (2).
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Setting the Sipper Safe Height and teaching the four touchpoints
e Close the doors to engage the motors. f Click Set and Next in the Sipper Configuration Wizard.
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Preparing to verify the calculated teachpoints
Preparing to verify the calculated teachpoints
To prepare to verify the calculated teachpoints:
Open the doors and install an empty sample plate on the stage. Ensure the plate is securely positioned, and then close the doors and proceed to "Accepting or editing the calculated teachpoints" on page 30.
Accepting or editing the calculated teachpoints
In this part of the Wizard, the calculated teachpoints for the corner wells, wash stations, and matrix bottles are displayed and you can accept the values or edit them. You will do the following:
Step Task
See...
1
View the calculated teachpoints for the corner "Viewing the calculated teachpoints for the
wells and accept or edit them.
corner wells" on page 31
2
View the calculated teachpoints for the wash "Viewing the calculated wash station
station teachpoints and accept or edit them. teachpoints" on page 32
3
View the calculated matrix teachpoints and
"Viewing the calculated matrix teachponts" on
accept or edit them.
page 33
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Accepting or editing the calculated teachpoints
Viewing the calculated teachpoints for the corner wells
To accept or edit the corner well teachpoints:
1 In the Set Plate Teachpoints area, view the calculated teachpoints for three corner wells of the plate. The teachpoints are calculated for the following corner wells, depending on the plate type you are teaching:
Plate size 96-well 384-well 1536-well
First corner well A1 A1 A1
Second corner well Third corner well
H1
A12
P1
A12
FF1
A48
2 Choose one of the following options:
· Accept these values and click Set and Next to continue to the calculated Wash Station teachpoints step of the wizard.
· Edit one or more of the values and when finished, close the doors, and click Set and then Next to continue to the calculated Wash Station teachpoints step of the wizard.
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NOTE
System preparation Accepting or editing the calculated teachpoints
In the following steps, the wizard will go to each of these teachpoints and you can verify or edit each teachpoint.
Viewing the calculated wash station teachpoints
To accept or edit the wash station teachpoints: 1 In the Set Wash Station Teachpoints area, view the teachpoints for the
Aqueous, Middle, and Organic wash stations.
NOTE
2 Choose one of the following options:
· Accept these values and click Set and Next to continue to the calculated matrix position teachpoints of the wizard.
· Edit one or more of the values and when finished, close the doors, and click Set and then Next to continue to the calculated matrix teachpoints of the wizard.
In the following steps, the wizard will go to each of these teachpoints and you can verify or edit each teachpoint.
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Accepting or editing the calculated teachpoints
Viewing the calculated matrix teachponts
To view or edit the matrix teachpoints: 1 In the Set Matrix Teachpoints area, view the calculated teachpoints for the
matrix bottle positions.
2 Choose one of the following options:
· Accept these values and click Set and Next to move the stage to the calculated x- and y-coordinates of the Aqueous Wash Station teachpoint.
· Edit one or more of the values and when finished, close the doors, and click Set and then Next to move the stage to the Aqueous Wash Station teachpoint.
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Verifying the calculated teachpoints
Verifying the calculated teachpoints
In this part of the wizard you verify or edit the calculated teachpoints. If you do not want to change a teachpoint, click Next to advance to the next step without changing the teachpoint. You will do the following:
Step Task 1 Verify the wash station teachpoints. 2 Verify the corner well teachpoints. 3 Verify the matrix position teachpoints.
See
"Verifying the wash station teachpoints" on page 35
"Verifying the corner well teachpoints" on page 38
"Verifying the matrix position teachpoints" on page 41
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Verifying the calculated teachpoints
Verifying the wash station teachpoints
To verify the wash station teachpoints:
1 Verify the Aqueous Wash Station teachpoint as follows:
a Ensure the sipper is situated over the center of the Aqueous Wash Station chimney in the x- and y-axes.
b To adjust, open the doors, and manually move the stage in the x- and y-axes to the correct location.
c Manually move the stage in the z-axis from its z-safe height to the calculated set value as shown below (1) (also shown in the Set Wash Station Teachpoint area in "Viewing the calculated wash station teachpoints" on page 32).
d Adjust the z-axis depth, if required.
The sipper tube should reach a depth sufficient to wash the outside of the sipper that is exposed to well samples during a run. The depth required depends on the well size and the sample volume.
e When the position is correct, close the doors, and click Set and Next to move the stage to the calculated x- and y-coordinates of the Organic Wash Station chimney teachpoint.
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Verifying the calculated teachpoints
2 Verify the Organic Wash Station teachpoint as follows:
a Ensure the sipper is situated over the center of the Organic Wash Station chimney in the x- and y-axes.
b To adjust, open the doors and manually move the stage in the x- and y-axes to the correct location.
c Manually move the stage in the z-axis from its z-safe height to the calculated set value as shown below (1) (also shown in the Set Wash Station Teachpoint area in "Viewing the calculated wash station teachpoints" on page 32).
d Adjust the z-axis depth, if required.
The sipper tube should reach a depth sufficient to wash the outside of the sipper that is exposed to well samples during a run. The depth required depends on the well size and the sample volume.
e When the position is correct, close the doors and click Set and Next to move the stage to the calculated x- and y- coordinates of the Middle Wash Station chimney teachpoint.
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Verifying the calculated teachpoints
3 Verify the Middle Wash Station teachpoint as follows:
a Ensure the sipper is situated over the center of the Middle Wash Station chimney in the x- and y-axes.
b To adjust, open the doors, and manually move the stage in the x- and y-axes to the correct location.
c Manually move the stage in the z-axis from its z-safe height to the calculated set value as shown below (1) (also shown in the Set Wash Station Teachpoints area in "Viewing the calculated wash station teachpoints" on page 32).
d Adjust the z-axis depth, if required.
The sipper tube should enter the middle wash station to a depth past the inner hole so that when the sipper purges, fluid will not escape.
e When the position is correct, close the doors, and click Set and Next to move the stage to the calculated x- and y-coordinates of the corner well teachpoint, H1 for a 96-well, P1 for a 384-well, and FF1 for a 1536-well plate.
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Verifying the calculated teachpoints
Verifying the corner well teachpoints
To verify the corner well teachpoints:
1 Verify the sipper teachpoint for the first corner well as follows:
a Ensure the sipper is situated over the center of the well in the x- and y-axes.
b To adjust, open the doors, and manually move the stage in the x- and y-axes to the correct location.
c Manually move the stage in the z-axis from its z-safe height to the calculated set value as shown below (1) (also shown in the Set Plate Teachpoints area in "Viewing the calculated teachpoints for the corner wells" on page 31).
d Adjust the z-axis depth, if required, and then close the doors.
The sipper tube should be fully immersed when sampling occurs.
e Click Set and Next to move the stage to the calculated x- and y-coordinates of the corner well teachpoint, A1.
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Verifying the calculated teachpoints
2 Verify the A1 well Position teachpoint:
a Ensure the sipper is situated over the center of the well in the x- and y-axes.
b To adjust, open the doors, and manually move the stage in the x- and y-axes to the correct location.
c Manually move the stage in the z-axis from its z-safe height to the calculated set value as shown below (1) (also shown in the Set Plate Teachpoints area in "Viewing the calculated teachpoints for the corner wells" on page 31).
d Adjust the z-axis depth, if required, and then close the doors. The sipper tube should be fully immersed when sampling occurs.
e Click Set and Next to move the stage to the calculated x- and y-coordinates of the third corner well teachpoint, A12, A24, or A48.
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Verifying the calculated teachpoints
3 Verify the third corner well teachpoint:
a Ensure the sipper is situated over the center of the well in the x- and y-axes.
b To adjust, open the doors, and manually move the stage in the x- and y-axes to the correct location.
c Manually move the stage in the z-axis from its z-safe height to the calculated set value as shown below (1) (also shown in the Set Plate Teachpoints area in "Viewing the calculated teachpoints for the corner wells" on page 31).
d Adjust the z-axis depth, if required, and then close the doors. The sipper tube should be fully immersed when sampling occurs.
e Click Set and Next to move the stage to the calculated x- and y-coordinates of the Matrix 1 Position teachpoint.
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Verifying the calculated teachpoints
Verifying the matrix position teachpoints
To verify the matrix position teachpoints:
1 Verify the Matrix 1 Position teachpoint:
a Ensure the sipper is situated over the center of the bottle in the x- and y-axes.
b To adjust, open the doors, and manually move the stage in the x- and y-axes to the correct location.
c Manually move the stage in the z-axis from its z-safe height to the calculated set value as shown below (1) (also shown in the Set Matrix Teachpoints area in "Viewing the calculated matrix teachponts" on page 33).
a Adjust the z-axis depth, if required, and then close the doors.
The sipper tube depth should be adequate to reach the maximum amount of sample.
b Click Next to move the stage to the calculated x- and y-coordinates of the Matrix Position 2 teachpoint.
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Verifying the calculated teachpoints
2 Verify the Matrix 2 Position teachpoint:
a Ensure the sipper is situated over the center of the bottle in the x- and y-axes.
b To adjust, open the doors, and manually move the stage in the x- and y-axes to the correct location.
c Manually move the stage in the z-axis from its z-safe height to the calculated set value as shown below (1) (also shown in the Set Matrix Teachpoints area in "Viewing the calculated matrix teachponts" on page 33).
d Adjust the z-axis depth, if required, and then close the doors. The sipper tube depth should be adequate to reach the maximum amount of sample.
e Click Next to move the stage to the calculated x- and y-coordinates of the Matrix Position 3 teachpoint.
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Verifying the calculated teachpoints
3 Verify the Matrix 3 Position teachpoint:
a Ensure the sipper is situated over the center of the bottle in the x- and y-axes.
b To adjust, open the doors, and manually move the stage in the x- and y-axes to the correct location.
c Manually move the stage in the z-axis from its z-safe height to the calculated set value as shown below (1) (also shown in the Set Matrix Teachpoints area in "Viewing the calculated matrix teachponts" on page 33).
d Adjust the z-axis depth, if required, and then close the doors. The sipper tube depth should be adequate to reach the maximum amount of sample.
e Click Next to move the stage to the calculated x- and y-coordinates of the Matrix Position 4 teachpoint.
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Verifying the calculated teachpoints
4 Verify the Matrix 4 Position teachpoint:
a Ensure the sipper is situated over the center of the bottle in the x- and y-axes.
b To adjust, open the doors, and manually move the stage in the x- and y-axes to the correct location.
c Manually move the stage in the z-axis from its z-safe height to the calculated set value as shown below (1) (also shown in the Set Matrix Teachpoints area in "Verifying the matrix position teachpoints" on page 41).
d Adjust the z-depth, if required, and then close the doors. The sipper tube depth should be adequate to reach the maximum amount of sample.
e Click Set and then close the Sipper Configuration Wizard.
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Checking the nanovalves
Checking the nanovalves
To check the nanovalves:
1 Click System Tools > Valve Tuner to open the Valve Configuration Utility dialog box.
2 Click Find next to each valve to home it. The Home status light turns green when the valve is homed.
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Checking the nanovalves
3 Click Inject and Load to change the status of the valves and to verify that the change in status is smooth and quiet.
The valve status lights are green when they are in the Load position and the values in the Cur field of the Positions area should be close to zero.
The valve status lights are blue when they are in the Inject position and the values in the Cur field of the Positions area should be high >1000.
If the status lights for a valve (V1, V2, V3) are yellow and the Home status light is red, click Find to home the valve.
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Preparing the peristaltic pump
Preparing the peristaltic pump
About the peristaltic pump
Your RapidFire 400 System has one of two types of peristaltic pumps: · ISM peristaltic pump whose flow rate is controlled by buttons on the device.
· L-100 peristaltic pump whose flow rate is controlled in the RapidFire software.
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Preparing the peristaltic pump
To prepare the peristaltic pump: 1 Ensure the pump is turned on. 2 Ensure the vacuum is on and at least -70 kPa.
3 In the RapidFire main software window, select the Pump 4 check box and click Apply to start the flow.
For ISM peristaltic pumps, the flow rate control is unavailable in the software.
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Preparing the peristaltic pump
For the L-100 peristaltic pumps, the flow rate sets to the default 10 RPM.
4 Verify the pump is running in the clockwise direction.
5 Set the pump flow rate:
a Ensure that fluid is moving through the tubing lines that connect Pump 4 to the wash station.
b For ISM pumps, adjust the pump flow by pressing the up/down arrow buttons on the pump control panel.
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Preparing the peristaltic pump
c For L-100 pumps, adjust the pump flow by entering a number in the RPM field and clicking Apply in the RapidFire main window software.
d When the fluid starts to flow into the wash station, adjust the flow rate until droplets of liquid adhere to the aqueous and organic wash inlet tubes for about 6-8 seconds.
6 Ensure that fluid is being aspirated from the center trough sufficiently and no overflow occurs on the wash station.
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Preparing the quaternary pumps
Preparing the quaternary pumps
This procedure primes the pumps and removes air bubbles in the fluidic lines.
To prepare the quaternary pumps:
1 Turn the diversionary valve (black knob) counterclockwise 90-180 degrees to open the valve.
2 In the RapidFire software window, select the check boxes for Pumps 1, 2, and 3, and then click Apply.
3 Verify that the composition for channel B, C, and D is 0%.
4 Set the flow rate for each pump to 5 mL/min, click Apply, and let the pumps operate for 2 minutes to prime channel A.
5 Change the solvent composition by entering 100% for B and 0% for A, C, and D, click Apply, and allow the pumps to operate for 2 minutes to prime channel B.
6 Change the solvent composition by entering 100% for C and 0% for A, B, and D, click Apply, and allow the pumps to operate for 2 minutes to prime channel C.
7 Change the solvent composition by entering 100% for D and 0% for A, B, and C click Apply, and allow the pumps to operate for 2 minutes to prime channel D.
8 Set the composition and flow rates to your assay-specific values, for example 100% A at 1.25 mL/min, and click Apply.
9 Turn the black knob clockwise until it is finger-tight to close the diversionary valve.
10 Check the pressure readings:
a Click System Tools > Valve Tuner to open the Valve Configuration Utility dialog box.
b Select the Inject position for valves 1, 2, and 3 and click System Tools > Pressure Graph, to check the pressure.
c Select the Load position for valves 1,2, and 3 and click System Tools > Pressure Graph, to check the pressure.
The maximum pressure is 20 MPa. The following table lists typical pressure values.
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Preparing the sipper
Table 4. Typical pressure values
Valve position V1, V2, and V3 Inject position
V1, V2, and V3 Load position
Pump number P1 P2 P3 P1 P2 P3
Back pressure ~1-10 MPa (10-100 bar) ~0.1-3 MPa ~1-10 MPa ~0.1-10 MPa (lower than inject) ~0.1-3 MPa (higher than inject) ~1-10 MPa (higher than inject)
If the value is outside the typical range shown for each pump at the load or inject position:
· Check the flow path for leaks (low pressure) or clogs (high pressure), for example, a plugged cartridge.
· Verify you are using proper and miscible solvents.
Preparing the sipper
This procedure uses pump 1 to flush the contents of the sipper and its tubing into the wash station drain.
To prepare the sipper:
In the RapidFire software window, click Flush Now in the Sipper Settings area.
The following occurs: a Pump 1 turns on. b The stage moves so that the sipper is positioned over the wash station drain. c Valve 4 actuates to the Load position. d The flow rate for pump 1 increases to 8 mL/min and maintains for one minute after which it returns to its previous setting. e Pump 1 turns off. f Valve 4 actuates to the Inject position. g The stage moves to the Swap position.
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Preparing the temperature control unit
Preparing the temperature control unit
To turn on the temperature control unit:
1 Select the Use Chiller check box if you want to maintain a temperature between 5-25 °C during the run.
2 In the Set pt(°C) field, enter the temperature that you want the unit to maintain during the run.
3 Click Apply to turn on the temperature control unit.
Preparing the TOF or Q-TOF mass spectrometer
Before you start
If you are using one computer to control both the RapidFire instrument and the mass spectrometer: · Ensure Mass Hunter Data Acquisition software 10.00.xx or later is installed on
the computer.
If you are using two computers, one to control the RapidFire instrument and one to control the mass spectrometer: · Ensure Mass Hunter Data Acquisition software 10.00.xx or later is installed on
the computer controlling the mass spectrometer. · Ensure RapidFire Communicator software version 6.0 or later is installed on
the computer controlling the mass spectrometer.
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Preparing the TOF or Q-TOF mass spectrometer
To prepare the TOF or Q-TOF mass spectrometer:
1 If you are using one computer to control the RapidFire instrument and the mass spectrometer, go to step 2.
If you are using two computers to control the RapidFire instrument and the mass spectrometer:
a Open RapidFire Communicator by double-clicking the desktop icon ( ).
b Open the Windows Task Manager and verify the following processes are running:
· RFRemServer.exe
· RFMasshunterS_TOF.exe
2 Start the Mass Hunter Data Acquisition software by double-clicking the Data Acquisition desktop icon.
3 To turn on the detector, click On in the Q-TOF device pane or in the status line for the Instrument Status area.
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The status indicator of the device in the Status area turns from teal (Standby, connected), to yellow (Not Ready, preparing), and then to green (Ready, idle).
4 Select Tune from the Context area.
5 Set the mass and resolution parameters for your analysis in the Instrument State tab in the Tune window.
Note that: · High Resolution (scan speed 4 GHz) · Extended Dynamic Range (scan speed 2 GHz) a Click Apply on the bottom right. b Wait for the system to equilibrate.
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Preparing the TOF or Q-TOF mass spectrometer
6 Calibrate the mass spectrometer: a Set the location of the Calibrant Bottle, for example B, and wait for the mass-over-charge spectrum to stabilize. b Click the Tune & Calibration tab. c Select the Polarity to use. You can select both, if desired. d Click Mass Calibration / Check. e Click Start TOF Mass Calibration.
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Preparing the TOF or Q-TOF mass spectrometer
f When the Check Tune Report appears, check the results. For example, check that the calculated residual errors are satisfactory (for example, <2 ppm).
NOTE
g Click OK to close the results window. h Set the Calibrant Bottle flag to None. 7 Select Acquisition from the Context list. If prompted, confirm that you want to operate under the modified Instrument State.
DO NOT modify the Tune File.
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Preparing the TOF or Q-TOF mass spectrometer
8 Click Method > Open to open a method (file type is .m) or set the parameters to create a method.
Methods for the RapidFire System are stored in the D:\MassHunter\methods\RapidFire folder.
9 Set the parameters for your analysis on the TOF or Q-TOF tabs. The following figures show example screens.
Figure 5. General tab example
Figure 6. Source tab example
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Preparing the TOF or Q-TOF mass spectrometer
Figure 7. Acquisition tab example
Figure 8. Reference Mass example
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Preparing the TOF or Q-TOF mass spectrometer
Figure 9. Chromatogram tab example
10 When you have finished setting the parameters, click the check mark ( ) in the toolbar area of the Method Editor window.
11 Save the method.
Save all method files for use with the RapidFire System in the D:\MassHunter\methods\RapidFire folder.
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Preparing the TQ or Ultivo TQ mass spectrometers
Preparing the TQ or Ultivo TQ mass spectrometers
Before you start
If you are using one computer to control both the RapidFire instrument and the mass spectrometer:
· Ensure Mass Hunter Data Acquisition software 10.00.xx or later is installed on the computer.
If you are using two computers, one to control the RapidFire instrument and one to control the mass spectrometer:
· Ensure Mass Hunter Data Acquisition software 10.00.xx or later is installed on the computer controlling the mass spectrometer.
· Ensure RapidFire Communicator software version 6.0 or later is installed on the computer controlling the mass spectrometer.
To prepare the TQ or Ultivo TQ mass spectrometer:
1 If you are using one computer to control the RapidFire instrument and the mass spectrometer, go to step 2.
If you are using two computers to control the RapidFire instrument and the mass spectrometer:
a Open RapidFire Communicator by double-clicking the desktop icon ( ).
b Open the Windows Task Manager and verify the following processes are running:
· RFRemServer.exe
· RFMasshunterS_TQ.exe
2 Start Mass Hunter Data Acquisition software by double-clicking the Data Acquisition desktop icon.
3 Turn on the detector by clicking On in the TQ device pane or in the status line for the Instrument Status area.
The status indicator of the device in the Status area turns from teal ((Standby, connected), to yellow (Not Ready, preparing), and then to green (Ready, idle).
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Minimizing the required sample volume
Minimizing the required sample volume
The volume used per sample well for the standard setup of the RapidFire 400 System is approximately 35 µL. This includes 10 µL of sample for measurement and 25 µL of dead volume, composed of a sample loop (A) and sipper tube (B).
It is possible to reduce the sample size to 5 µL and 10 µL with the use of smaller diameter sipper and sample loop tubing. The use of 1536-well microplates require smaller sample size because of the limited volume in each well.
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Minimizing the required sample volume
The smaller diameter tubing and other required parts are available from Agilent Technologies. The required parts and the amount of sample that is measured are listed in the table below.
Parts for 10 µL sample measurement
Part number RF0052T RF0111T-8
RF0094T
Quantity 1 1
1 (10 pk)
Description
Notes
0.5" guide needle
Replaces 1.5" guide needle
8" OF 0.009" ID tubing, gray Cut to length (4.5" for sipper tube, 3.5" for sample loop)
Ferrule, red od 1/32"
One per connection
Parts for 5 µL sample measurement
Part number RF0052T RF0112T-13
RF0094T
Quantity 1 1
1 (10 pk)
Description 0.5" guide needle 12" of 0.005" id tubing, red
Ferrule, red od 1/32"
Notes
Replaces 1.5" guide needle
Cut to length (4.5" for sipper tube, 3.5" for sample loop)
One per connection
To run the system with 5 µL and 10 µL sample volumes:
1 Prepare the RapidFire 400 system:
a Replace the standard guide needle with the 0.5" guide needle.
b Replace the standard sample loop tubing and sipper tubing with the desired smaller diameter loop and sipper tubing.
2 To account for the shorter guide needle, reteach the stage teachpoints using the appropriate plate type. For details, see "Setting the stage teachpoints" on page 18.
3 Using a standard analyte, empirically determine the optimal sip time for your system by inactivating the sip sensor and creating sample runs with sip times between 50-200 ms. The optimized sip time will be used for the Aspirate step of the RapidFire method.
For more information on performing runs with minimal sample volumes, contact customer support at Agilent Technologies.
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Shutting down the RapidFire system between runs
Shutting down the RapidFire system between runs
NOTE
When you have completed your runs using the RapidFire system, shut down the system as follows.
The instrument goes into stand-by mode at the end of each run. This ensures that all pumps are turned off at the end of a run.
To shut down the system: 1 If your samples contain high-salt and/or high-protein:
a Flush the sipper to avoid the formation of precipitates as described in "Preparing the sipper" on page 52.
b Clean the sample flow path by running a minimum of 50 wells containing methanol.
2 Close the RapidFire Software: a In the RapidFire software window, click Disconnect to disconnect the software from the instrument. b Close the RapidFire software window. c In the RapidFire Control Panel, click Stop. d Close the RapidFire Control Panel.
3 Put the mass spectrometer into standby mode. 4 Turn off the RapidFire vacuum source. DO NOT turn off the mass
spectrometer vacuum source.
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Operating the RapidFire 400 in Plates mode
This section describes how to operate the RapidFire 400 in Plates mode. It contains the following topics: About Plates mode 66 Workflow for using Plates Mode 66 Before you start 66 Recommendations for preparing the plates 67 Loading a single plate 68 Loading multiple plates using the plate handler 68 Preparing the TOF or Q-TOF mass spectrometer 70 Preparing the TQ or Ultivo TQ mass spectrometer 72 Setting up the RapidFire parameters for a Plates mode run 74 Starting a run in Plates mode 78 Pausing a run in Plates mode 79 Stopping a run in Plates mode 79 Maintaining a log book of data 80
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About Plates mode
About Plates mode
Plates mode can be used when you have one or more plates that you want to sample without a preset method that has a specific sipping sequence.
In Plates mode, you load your plate or plates and run them according to the conditions chosen in the RapidFire software window. The samples are fed into the mass spectrometer but the data collection is not synchronized to the RapidFire run.
This mode is primarily used for testing and optimizing conditions for data collection when using Sequence mode.
Workflow for using Plates Mode
Table 5. Plates Mode workflow steps
Step Task
See...
1
Prepare the system
"Before you start" on page 66
1
Prepare plates
"Recommendations for preparing the plates" on page 67
2
Load plates
· "Loading a single plate" on page 68 · "Loading multiple plates using the plate handler" on page 68
3
Prepare MS
· "Preparing the TOF or Q-TOF mass spectrometer" on page 70 · "Preparing the TQ or Ultivo TQ mass spectrometer" on page 72
4
Set RF run parameters "Setting up the RapidFire parameters for a Plates mode run" on
page 74
5
Start run
"Starting a run in Plates mode" on page 78
Before you start
Ensure you have prepared the RapidFire system. Refer to the following topics: · "Starting the system" on page 13 · "Loading and homing the cartridge changer" on page 16
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Recommendations for preparing the plates
· "Minimizing the required sample volume" on page 62, if you are running 1536-well plates or minimizing sample volume
· "Homing the stage" on page 18 · "Checking the nanovalves" on page 45 · "Preparing the peristaltic pump" on page 47 · "Preparing the quaternary pumps" on page 51 · "Preparing the sipper" on page 52. · "Preparing the temperature control unit" on page 53
Recommendations for preparing the plates
· Ensure sample plates have thawed completely. · Centrifuge the plates at 3000 rpm for at least five minutes.
Centrifuging the plates forces particles and insoluble material to the bottom of the wells where they are less likely to be aspirated into the system.
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Operating the RapidFire 400 in Plates mode
Loading a single plate
Loading a single plate
To load a single plate:
1 Ensure that the stage has been homed. See "Homing the stage" on page 18 for a procedure.
2 Open the doors and securely position the plate on the stage. Ensure well A1 is aligned with the A1 marking on the stage.
3 Close the doors.
Loading multiple plates using the plate handler
Before you start:
If you are using the temperature control unit, make sure it is on and has reached the set temperature. See step 9 in "Setting up the RapidFire parameters for a Plates mode run" on page 74.
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Loading multiple plates using the plate handler
To load plates using the plate handler: 1 In the RapidFire software window:
a Select the plate type you are using from the Plate Configuration list. b Select Use Plate Handler and click Apply.
2 Open the doors and pull out the drawer slide containing the three plate stacks.
3 Load the plates into two of the stacks. The A1 well should be oriented on the right-hand side towards the back of the unit as you are standing in front of the system and looking towards the back.
Make sure one stack is empty so it can receive plates once they have been sampled.
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Preparing the TOF or Q-TOF mass spectrometer
Preparing the TOF or Q-TOF mass spectrometer
To prepare the TOF or Q-TOF mass spectrometer:
1 Set up the mass spectrometer as described in "Preparing the TOF or Q-TOF mass spectrometer" on page 53.
2 In the Sample Run window of the MassHunter Acquisition software:
a Enter the Name and Path for the data file.
The data file name must be sequence1.d (for data analysis by RapidFire Integrator).
b Click to add a parameter to the Additional Information list.
c Select RunType and click OK.
d For the RunType, select Manual Run.
e Set the Injection Volume to As Instrument. This value causes the system to use the injection volume set by the RapidFire injection loop (line connected between port 2 and 5 on valve 1).
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Preparing the TOF or Q-TOF mass spectrometer
3 Click the start arrow ( ) to start the run.
When the source has equilibrated, the statue indicator of the TOF or Q-TOF device module turns blue in the Instrument Status area.
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Preparing the TQ or Ultivo TQ mass spectrometer
Preparing the TQ or Ultivo TQ mass spectrometer
To prepare the TQ mass spectrometer:
1 Set up the mass spectrometer as described in "Preparing the TQ or Ultivo TQ mass spectrometers" on page 61.
2 Select Acquisition from the Context list.
3 To open a .m method or set parameters for a new method, click Method > Open.
Agilent recommends storing methods for the RapidFire system in the D:\MassHunter\methods\RapidFire folder.
4 Set the parameters for your analysis on the TQ tabs.
5 In the Sample Run window in the MassHunter Data Acquisition software:
a Enter the Name and Path for the data file.
The data file Name must be sequence1.d (for data analysis by RapidFire Integrator software).
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Preparing the TQ or Ultivo TQ mass spectrometer
b Click to add a parameter to the Additional Information list.
c Select RunType and click OK.
d For the RunType, select Manual Run.
e Set the Injection Volume to As Instrument. This value causes the system to use the injection volume set by the RapidFire injection loop (at V1P2-V1P5).
6 Click Start Sample Run, or click Sample > Run.
When the source has equilibrated, the status indicator of the TQ device module turns blue in the Instrument Status area.
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Setting up the RapidFire parameters for a Plates mode run
Setting up the RapidFire parameters for a Plates mode run
Before you start
Make sure that the cartridges are installed and the Cartridge Changer has been homed. See "Loading and homing the cartridge changer" on page 16 for a procedure.
To set up the RapidFire run parameters for Plates mode: 1 In the RapidFire software window, click System Tools > Cartridge Changer.
The Cartridge Changer dialog box opens. 2 Select the position you want to use from the Go To list and click Go To.
Ensure the changer moves to the correct position the field to the right of the drop list should display the same number as shown in the Go To list.
3 Close the Cartridge Changer dialog box.
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Setting up the RapidFire parameters for a Plates mode run
4 In the RapidFire software window, select Plates in the Plateset Run Controls area.
5 In the Plates Mode Settings area, set the following:
a Type the position of the well on the plate to start the run in Start at Well fields. The first position is A1.
b Select the order in which you want the run to proceed,
· By Row (for example, A1, A2, A3...H11, H12)
· By Col (for example, A1, B1, C1...G12, H12)
6 In the Sipper Settings area, set the following:
a Select the Wash Needle Between Sips check box to wash the sipper once after aspiration of the sample from each well. The sipper is washed first in the aqueous, and then in the organic wash chimney for 300 ms each.
b In the Sip Height field, enter the number of millimeters (1 mm or more) from the lowest position of the sipper to the bottom of the well.
The sipper (in valve 1, port 3) travels down from the safe z position into the sample well until either:
· The sip sensor (in valce 1, port 4) optically detects the presence of fluid and trigger the actuation of valve 1, which causes the transition between State #1 (Aspiration) and State #2 (Load/Wash), or
· The z-position of the sipper reaches the set Sip Height.
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Setting up the RapidFire parameters for a Plates mode run
c If you want the system to do extra cycles in the organic wash station between samplings, enter a value (more than 0) for Blank Injections Between Wells.
This setting helps to limit or prevent carryover between samples.
7 In the RapidFire Cycle Durations area, set the valve timing as per assay protocol for Aspirate, Load/Wash, Extra Wash, Elute, and Reequilibrate.
Set Aspirate to at least 600 ms for reliable sip sensor triggering.
8 In the Plate Info area, select the plate type you are using from the Plate Configuration list.
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Setting up the RapidFire parameters for a Plates mode run
9 In the Miscellaneous Settings area, do the following:
a Select the Use Plate Handler check box if you are using the plate handler to deliver the plates.
b Select the Use Barcode Scanner check box if you want to scan the plate barcode before the plate is loaded onto the stage.
c Do not select the Mass Spec Standby After Run check box. This option is not available in Plates mode.
d In the Plates Between Flushes field, enter the number of plates that should be sampled before the sipper is flushed for 1 min. Recommended values are 4 for a 96-well plate and 1 for a 384-well plate, but will depend on the specifics of a given method.
e In the Missed Sips Tolerance field, enter the number of times the sip sensor can miss detection of a sip before the system stops operation.
f Systems with a temperature control unit. Select the Use Chiller check box if you want to maintain a temperature below ambient during the run. Enter the temperature (°C) in the Set pt field that you want the unit to maintain during the run.
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Starting a run in Plates mode
10 In the Pump Settings area do the following: a Set pump flow rates for your run. b Set the solvent compositions for your run. c Set the minimum and maximum pressure for each pump.
11 Click Apply.
12 To save the parameter settings for future use, click File > Save RF Method As and enter a name for the method and click Save.
Starting a run in Plates mode
Before you start
· Initiate the mass spectrometer acquisition run before you start the RapidFire run. (Runs in Plates mode are not synchronized with the mass spectrometer.)
· Make sure the doors are closed and the emergency stop is in the released position.
To start a run in Plates mode:
1 Click start
in the upper left area of the screen.
2 When prompted, enter the plate name or input the barcode using the hand-held barcode scanner and then click OK.
3 Verify that the sipper is aspirating the samples and the analytes are being detected by the mass spectrometer.
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Pausing a run in Plates mode
4 If you are loading plates manually, when the plate run is complete, you will be prompted to load a new plate. Do one of the following:
a If you have more plates to run:
· Open the doors and remove the previous plate.
· Place your next plate on the stage as described in "Loading a single plate" on page 68 and close the doors.
· Enter a name for the new plate or scan the plate barcode with the hand-held barcode scanner.
If you do not enter a plate name within 30 minutes, a timeout automatically puts the RapidFire System in standby mode to prevent wasting solvent.
b If you are done running plates, click Cancel.
Pausing a run in Plates mode
To pause a run:
1 Click pause .
The system pauses after the cycle of Aspirate, Load/Wash, Extra Wash, Re-equilibrate, and Needle Wash completes. The mass spectrometer continues to collect baseline data and the RapidFire pumps continue to run.
2 When you are ready to resume operation, click start
.
The system resumes operation where it left off and data is saved to the current data file.
Stopping a run in Plates mode
To stop a run that you do not want to restart again: 1 Click stop .
The system stops after the cycle of Aspirate, Load/Wash, Extra Wash, Re-equilibrate, and Needle Wash completes. Data acquisition stops and the current data file closes.
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Maintaining a log book of data
Maintaining a log book of data
Agilent recommends keeping a record of the data sets that are stored in the computer in order to retrieve the data more easily and match the Plates mode data sets of the RapidFire system and mass spectrometer in data analysis.
Consider including the following information in the record book, such as in a spreadsheet format:
· Date Generated data is stored by date in the RapidFire computer. See the Data Analysis Guide for more information.
· Assay description What RF and MS methods were used.
· Sample name What the RF and MS data file names were used.
· Set of plates number as assigned by the RapidFire system. See the Data Analysis Guide for more information.
· Plate number (in the current set of plates)
· Comments
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mode
This chapter describes how to operate the RapidFire 400 system in Sequences mode. It contains the following topics: About Sequences mode 82 Workflow for operating in Sequences mode 83 Before you start 84 Creating a RapidFire Method file (.rfcfg) 84 Creating a RapidFire Sequence PlateMap file (.rfcfg) 88 Creating a RapidFire Batch file 90 Recommendations for preparing the sample plates 95 Loading a single plate 95 Loading multiple plates 96 Preparing the TOF or Q-TOF mass spectrometer 97 Preparing the TQ or Ultivo TQ mass spectrometer 98 Starting a run in Sequences mode 98 Using an information management file for a Sequences mode run 99 Creating a map file using MapFile Generator utility 100 Starting the run in Sequences mode using an information management file 108 Pausing a run in Sequences mode 110 Stopping a run in Sequences mode 111 Maintaining a log book of data 111
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About Sequences mode
About Sequences mode
Sequences mode can be used when you want to assay one or more sequences from sample wells in one or more plates. The plate or plates are run as a batch and the data collection by the mass spectrometer is synchronized with the RapidFire sample processing.
The following figure shows the structure of a batch and the files that make up a batch.
A batch file (.bat) consists of one or more plates. Each plate can have one or more sequences to be sampled. Each sequence is associated with a plate map (.rfmap) that defines the wells to sample. Each plate map is associated with an RF method (.rfcfg) and cartridge. The sample exiting the cartridge is ported directly to the mass spectrometer which analyzes the sample according to a preselected mass spectrometer method.
Figure 10. Structure of a batch file.
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Workflow for operating in Sequences mode
Workflow for operating in Sequences mode
Table 6. Workflow for Sequences mode
Step
Task
1
Prepare the system
1
Create a method file
2
Create plate map file
3
Create batch file
4
Prepare plates
5
Load plates
6
Prepare MS
7
Start the run
8
Pause the run
9
Stop the run
See
"Before you start" on page 84
"Creating a RapidFire Sequence PlateMap file (.rfcfg)" on page 88
"Creating a RapidFire Sequence PlateMap file (.rfcfg)" on page 88
"Creating a RapidFire Batch file" on page 90
"Recommendations for preparing the sample plates" on page 95
· "Loading a single plate" on page 95 · "Loading multiple plates" on page 96
· "Preparing the TOF or Q-TOF mass spectrometer" on page 97
· "Preparing the TQ or Ultivo TQ mass spectrometer" on page 98
"Starting a run in Sequences mode" on page 98
"Pausing a run in Sequences mode" on page 110
"Stopping a run in Sequences mode" on page 111
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Before you start
Before you start
Ensure you have prepared the RapidFire system. Refer to the following topics: · "Starting the system" on page 13 · "Loading and homing the cartridge changer" on page 16 · "Minimizing the required sample volume" on page 62, if you are running
1536-well plates or minimizing sample volumes · "Homing the stage" on page 18 · "Checking the nanovalves" on page 45 · "Preparing the peristaltic pump" on page 47 · "Preparing the quaternary pumps" on page 51 · "Preparing the sipper" on page 52 · "Preparing the temperature control unit" on page 53
Creating a RapidFire Method file (.rfcfg)
Running the RapidFire in Sequences mode requires a RapidFire Method file (.rfcfg). You create the Method file by setting the parameters in the RapidFire UI main window and then saving them as an .rfcfg file. The Method file saves the parameter settings in the following areas: · Sipper Settings · RapidFire Cycle Durations · Pump Settings
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Creating a RapidFire Method file (.rfcfg)
To set the parameters for the method:
1 In the Sipper Settings area, set the following:
a Select the Wash Needle Between Sips check box to wash the sipper once after aspiration of the sample from each well. The sipper is washed first in the aqueous, and then in the organic wash chimney for 300 ms each.
b In the Sip Height field, enter the number of millimeters (1 mm or more) from the lowest position of the sipper to the bottom of the well.
The sipper (in valve 1, port 3) travels down from the safe z position into the sample well until either:
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Creating a RapidFire Method file (.rfcfg)
· The sip sensor (in valve 1, port 4) optically detects the presence of fluid and triggers the actuation of valve 1, which causes the transition between State #1 (Aspiration) and State #2 (Load/Wash).
· The z-position of the sipper reaches the set Sip Height.
c If you want the system to do extra cycles in the organic wash station between samplings, enter a value (more than 0) for Blank Injections Between Wells.
This setting helps to limit or prevent carryover between samples.
2 In the RapidFire Cycle Durations area, set the valve timing as per assay protocol for Aspirate, Load/Wash, Extra Wash, Elute, and Reequilibrate.
Set Aspirate to at least 600 ms for reliable sip sensor triggering.
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Creating a RapidFire Method file (.rfcfg)
3 In the Pump Settings area do the following: a Set pump flow rates for your run. b Set the solvent compositions for your run. c Set the minimum and maximum pressure for each pump.
4 Click Apply.
5 Click File > Save RF Method As, enter a name for the method and click Save.
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Creating a RapidFire Sequence PlateMap file (.rfcfg)
Creating a RapidFire Sequence PlateMap file (.rfcfg)
NOTE
Running the RapidFire system in Sequences mode requires a RapidFire Sequence PlateMap file (.rfmap).
To create a RapidFire Sequence Platemap file:
1 In the RapidFire UI window, click Batch Configuration Tools > Plate Map Editor.
2 Select a Plate Size, 96-, 384-, or 1536-wells.
3 Create a Sequence:
a Click New.
The number 1 appears in the Sequences field indicating this is the first sequence.
b Click on a well, Matrix Station (1-4), or Wash Station (Aqueous, Organic) to add it to the sequence. To select multiple wells, click and drag the cursor over the wells you want to select.
c View the sequence in the Current Sequence box. You can edit the sequence in this area.
d To create another sequence, repeat this step.
If you are running 1536-well plates with a TOF or QTOF mass spectrometer and acquiring large amounts of data by scanning at a slow rate or a wide mass range, the data file might become too large for the software to parse and analyze. To avoid this, you can create multiple sequences within a plate so that the data will be spread among multiple files.
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Creating a RapidFire Sequence PlateMap file (.rfcfg)
4 Optional. You can enter a special well designation for each sequence to insert a pause in the RapidFire system data acquisition, as follows:
a Place the cursor in the Current Sequence field at the point you want to pause, for example, at the end of a sequence.
b Enter the time (in seconds) you want to delay the acquisition in the field next to Delay.
c Click Delay.
5 Optional. You can enter a flush at a desired point in the sequence, for example, between sequences, as follows:
a Place the cursor in the Current Sequence field at the point you want to flush.
b Enter the time (in seconds) you want to flush the sipper in the field next to Flush.
c Click Flush.
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Creating a RapidFire Batch file
6 Click File > Save As. Enter a name for the Sequence PlateMap (.rfmap) and click Save.
Creating a RapidFire Batch file
After you have created a method and plate map for the RapidFire system, you can create a batch file. To create a batch file: 1 In the RapidFire software window, click Batch Configuration Tools > Batch
Editor. The Batch Editor dialog box opens. 2 Select the mass spec you are using from the MS Interface list.
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Creating a RapidFire Batch file
3 Click Add.
The Open Sequence File dialog box opens.
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Creating a RapidFire Batch file
4 Select the sequence file (.rfmap) you want to use and click Open.
5 In the New Plate dialog box, enter the barcode or name of the plate that will be processed and click OK.
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The Batch Editor dialog box displays the sequences identified in the plate map.
6 For each sequence, double-click in the RF Method field to open the Open File dialog box, select the RapidFire method (.rfcfg) file you want to use, and click Open.
7 For each sequence, in the Cartridge field, select the cartridge to use from the list.
8 For each sequence, in the MS Method field, select the MS Method to use from the list. (The methods should be located in D:\MassHunter\methods\RapidFire.)
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Creating a RapidFire Batch file
9 Click File > Save As.
10 Enter a name for the batch file and click Save.
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Recommendations for preparing the sample plates
Recommendations for preparing the sample plates
· Completely thaw the sample plates. · Centrifuge the plates at 3000 rpm for at least five minutes.
Centrifuging the plates forces particles and insoluble material to the bottom of the wells where they are less likely to be aspirated into the system.
Loading a single plate
To load a single plate: 1 Ensure that the stage has been homed. See "Homing the stage" on page 18. 2 Open the doors and securely position the plate on the stage.
3 Close the doors.
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Loading multiple plates
Loading multiple plates
Before you start
If you are using the temperature control unit, make sure the unit is on and the unit has reached the set temperature.
To load multiple plates: 1 In the RapidFire software window:
a Select the plate type you are using from the Plate Configuration list. b Select Use Plate Handler and click Apply.
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Preparing the TOF or Q-TOF mass spectrometer
2 Open the doors and pull out the slide drawer containing the three plate stacks.
3 Load the plates into two of the stacks. The A1 well should be oriented on the right-hand side towards the back of the unit as you are standing in front of the system and looking towards the back.
Make sure one stack is empty so it can receive plates once they have been sampled.
4 Close the drawer.
Preparing the TOF or Q-TOF mass spectrometer
To prepare the TOF or Q-TOF mass spectrometer: 1 Set up the mass spectrometer as described in "Preparing the TOF or Q-TOF
mass spectrometer" on page 53. 2 Check that the Instrument Status is Ready (green) in the MassHunter Data
Acquisition software.
In Sequences mode, the RapidFire system and MS system are synchronized and the data is recorded automatically in D\MassHunter\Data\RapidFire. Refer to the Data Analysis Guide for more information.
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Preparing the TQ or Ultivo TQ mass spectrometer
Preparing the TQ or Ultivo TQ mass spectrometer
To prepare the TQ mass spectrometer:
1 Set up the mass spectrometer as described in "Preparing the TQ or Ultivo TQ mass spectrometers" on page 61.
2 Confirm that all MS methods used in Sequences mode operation in RapidFire are present in the folder D:\MassHunter\methods\RapidFire, as required for Agilent MS integration.
Starting a run in Sequences mode
Before you start
1 In the RapidFire UI window, click File > Load RF batch and select the batch file (.rfbat) you want to run and click Open.
2 Set the parameters in the Miscellaneous Settings area: a Select the Use Barcode Scanner check box if you are using the system barcode scanner. b In the Plates Between Flushes field, enter the number of plates to run before flushing the system. c In the Missed Sip Tolerance field, enter the number of missed sips to allow before triggering an error. d Click Apply at the bottom of the window.
3 Make sure the doors are closed and the emergency stop is in the released position.
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Using an information management file for a Sequences mode run
To start the run in Sequences mode:
1 Click start
in the upper left area of the screen.
2 When prompted, enter the plate name or input the barcode using the hand-held barcode scanner and then click OK.
3 Verify that the sipper is aspirating the samples and the analytes are being detected by the mass spectrometer.
4 If you are loading plates manually, when the plate run is complete, you will be prompted to load a new plate. Do one of the following:
a If you have more plates to run:
· Open the doors and remove the previous plate.
· Place your next plate on the stage as described in "Loading a single plate" on page 95 and close the doors.
· Enter a name for the new plate or scan the plate's barcode with the hand-held barcode scanner.
If you do not enter a plate name within 30 minutes, a time-out automatically puts the RapidFire System in standby mode to prevent wasting solvent.
b If you are done running plates, click Cancel.
Using an information management file for a Sequences mode run
You can use the MapFile Generator Utility to map your sample data columns from an information management file, for example LIMS, to a batch file (.bat) created in the RapidFire software. Once you create a map file, you import your sample data file before running a batch file.
Workflow
Step For this task...
1
Create a batch file
2
Create a map file
3
Start the run
See...
"Creating a RapidFire Batch file" on page 90
"Creating a map file using MapFile Generator utility" on page 100
"Starting the run in Sequences mode using an information management file" on page 108
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Creating a map file using MapFile Generator utility
Creating a map file using MapFile Generator utility
To create a map file: 1 In the RapidFire UI window, click File > Generate Map file.
The Map File Generator dialog box opens.
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2 Click the browse button ( ) next to the Example Import File Path field and select the location of the folder containing the sample file you wish to map.
Once selected, the files within this folder are accessible from the Example Import File field.
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Creating a map file using MapFile Generator utility
3 Select the specific file you want to map from the Example Import File list.
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Creating a map file using MapFile Generator utility
The Select Worksheet field displays the file type if the Example Import File is either a .txt or .csv file type. If the Example Import File is an Excel file (.xlsx), then the Select Worksheet field displays all the sheets in the Excel file.
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Creating a map file using MapFile Generator utility
4 Click Add All Columns to display the column information from the input file.
5 Map the headings of the input column to the RapidFire column names: a Select System for the Column Type for all input columns.
b Select a heading from the Map Column list that best corresponds to the heading in the input file.
For the import sample data feature to work correctly, make sure:
· The input data file has columns containing information of Sample Position and Barcode and the Sample Position is in the format A1, A2 and so forth.
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· The index of columns in the sample input file for Sample Position, Barcode, and Run Template match the index of the corresponding fields in the map file.
For example, if the input file has the Plate Position as the eighth column, then the Plate Position column must also be in the eighth position in the map file.
6 When you have matched all the input columns of interest, you can delete the ones not used.
To delete a column header:
a Select the row containing the column header.
b Right-click and select Delete.
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Creating a map file using MapFile Generator utility
7 Set the data start line for the input file: a Click Settings at the top of the window.
b In the First lines as comments field, enter the number of lines that occur before the start of the actual data, including the header line, but not any comment lines.
For example, if you have no comment lines and no -extra- lines before the column header line, then enter 1 in this field.This represents the one line describing the column headers.
8 Click Home and then click Save and select Save As to open the Save file dialog box.
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9 Select a location to save the file, type a name in the File name field, ensure the Save as type is Map Files.xml, and click Save.
10 Close the Map File Generator utility.
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Starting the run in Sequences mode using an information management file
Starting the run in Sequences mode using an information management file
To start the run:
1 In the RapidFire UI main window, click File > Load RF Batch and open the batch file you want to run using the sample data file you mapped in the previous procedure.
2 Click File > Import Sample Data. The Import Sample Data dialog box opens.
3 Fill in the data fields according to the following table:
Table 7. Import Sample Data dialog box parameters:
Parameter Operator Name Sample Path
Description/Action
Optional. Type in a name.
Source file to be analyzed. Click the Browse button and select the sample input file you used in the mapping procedure.
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Table 7. Import Sample Data dialog box parameters:
Parameter Map Path
Append New Data Directory Name
Injection name suffix
Description/Action
Map file to use to match the columns of the sample input file to the Mass Hunter Quant columns. Click the Browse button and select the file.
Select this option if this sample import is a continuation of a series of plates
Select this option if this is the first imported sample data file
Name of the directory where the MS data files will be stored. If left blank, the name will follow the convention year/month/day/number of plates analyzed.
Suffix added to the data file name. To activate the list, click Start Import and then select the desired suffix from the list.
4 If you have additional plate files, repeat step 3 and be sure you select the Append option.
The Import Log area lists all the wells for each file of the imported plate in the order in which they were imported.
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NOTE
Operating the RapidFire 400 in Sequences mode Pausing a run in Sequences mode
5 Click Close and start the run by clicking the start button ( ).
Once the data is acquired and converted, the sample information will be retained when opening the data in MassHunter Analysis software.
6 When the run is finished, in the RapidFire software window, click File > Clear Sample Data before starting a new import sequence.
Pausing a run in Sequences mode
To pause a run in Sequences mode:
1 Click pause .
The system pauses after the cycle of Aspirate, Load/Wash, Extra Wash, Reequilibrate, and Needle Wash completes. The mass spectrometer continues to collect baseline data and the RapidFire pumps continue to run.
2 When you are ready to resume operation, click start
.
The system resumes operation where it left off and data is saved to the current data file.
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Stopping a run in Sequences mode
Stopping a run in Sequences mode
To stop a run in Sequences mode:
1 Click stop .
The system stops after the cycle of Aspirate, Load/Wash, Extra Wash, Re-equilibrate, and Needle Wash completes. Data acquisition stops and the current data file closes.
Maintaining a log book of data
Agilent recommends keeping a record of the data sets that are stored in the computer in order to retrieve the data more easily and match the data sets of the RapidFire and mass spectrometer in Data Analysis.
Consider including the following information in the record book, such as in a spreadsheet format: · Date
Generated data is stored by date in the RapidFire computer. See the Data Analysis Guide for more information. · Assay description What RF and MS methods were used. · Sample name What the RF and MS data file names were used. · Set of plates number as assigned by the RapidFire system. See the Data Analysis Guide for more information. · Plate number (in the current set of plates) · Comments
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Maintenance
This section describes the maintenance requirements for the RapidFire 400 System and how to shut the system down. This section contains the following topics:
Daily maintenance 114
Weekly maintenance 114
Shutting down the RapidFire system completely 114
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Daily maintenance
Daily maintenance
To maintain the system daily:
1 Flush the sipper as described in "Preparing the sipper" on page 52.
2 Do the shut down procedures described in "Shutting down the RapidFire system completely" on page 114.
Weekly maintenance
To maintain the system weekly: 1 Empty the waste containers, carboy and vacuum waste container in
accordance with good laboratory practices and local, state/provincial, or national environmental and health regulations. 2 Clean the mass spectrometer according to the documentation for the mass spectrometer.
Shutting down the RapidFire system completely
To shut down the system completely: 3 Click Disconnect in the RapidFire software main window to disconnect the
software from the instrument. 4 Close the RapidFire software main window. 5 Click Stop in the RapidFire Control Panel. 6 Close the RapidFire Control Panel. 7 If the RapidFire system and the mass spectrometer are being controlled by
one computer, close the MassHunter Acquisition software. 8 Turn off the system by pressing the power switches on the back of the
RapidFire system (NOT the power to the mass spectrometer). 9 Turn off the vacuum source for the RapidFire system (NOT the vacuum for
the mass spectrometer).
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Shutting down the RapidFire system completely
10 Follow the mass spectrometer documentation instructions for shutting down the mass spectrometer.
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RapidFire System flow path reference
This sections provides figures that describes the flow paths for the various states of the nanovalves. This section contains the following topics: State 1: Aspirate flow path 118 State 2: Wash/Load flow path 119 State 3: Extra Wash flow path 120 State 4: Elute flow path 121 State 5: Re-equilibrate flow path 122 Flush the sipper flow path 123 Physical colors of the tubing 124 Valve positions 124
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State 1: Aspirate flow path
State 1: Aspirate flow path
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State 2: Wash/Load flow path
State 2: Wash/Load flow path
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State 3: Extra Wash flow path
State 3: Extra Wash flow path
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State 4: Elute flow path
State 4: Elute flow path
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State 5: Re-equilibrate flow path
State 5: Re-equilibrate flow path
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Flush the sipper flow path
Flush the sipper flow path
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Physical colors of the tubing
Physical colors of the tubing
Valve positions
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Changing the sequence of the sipper operation
This section provides a description of the change in the default sip sequence operation in RapidFire Acquisition software version 6.1 and how to implement this change if you are running RapidFire Acquisition software version 6.0 or earlier. This section contains the following topics:
About the sip sequence operation 126
Changing the sequence of sip operation 127
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Changing the sequence of the sipper operation
About the sip sequence operation
About the sip sequence operation
In RapidFire Acquisition software 6.1, the sequence of sip operation was changed to decrease the timing variability between well sampling. As a consequence, this change also slightly increases the through-put of the system.
In RapidFire Acquisition software version 6.0 and earlier, the default sequence of sip operation is as follows:
1 Draw Sample 1
2 Move to Wash Stations
3 Wait for current injection to conclude
4 Move to next sample location
5 Draw Sample 2
In RapidFire Acquisition software 6.1, the default sequence of sip operation was changed to the following:
1 Draw Sample 1
2 Move to Wash Stations
3 Move to next sample location
4 Wait for current injection to conclude
5 Draw Sample 2
If you have RapdiFire Acquisition software 6.0, or earlier, you can change the default sequence of sip operation to the current (6.1 version) default. See the following procedure "Changing the sequence of sip operation" on page 127 for how to make this change.
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Changing the sequence of the sipper operation
Changing the sequence of sip operation
Changing the sequence of sip operation
This procedure is for users upgrading to RapidFire Acquisition software 6.1 from version 6.0 or earlier.If this is a new RapidFire Acquisition software 6.1 installation, this change is included in the default cfg files.
Before you start
Close the RapidFire Control Panel and User Interface software, if running.
To change the sequence of sip operation: 1 Open the Sipper.cfg file in Notepad or other text editor. This file is located in
C:\agilent\RapidFire\Cfgs(6.0.0) #SGxxxxxx (where #SGxxxxxx is the serial number of your system). 2 Scroll to the bottom of the file and add the following line of text:
POSITION_OVER_NEXT_WELL_DURING_SIP_CYCLE=1;
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Changing the sequence of the sipper operation
Changing the sequence of sip operation
3 Save and close the file.
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Changing the sequence of the sipper operation
Changing the sequence of sip operation
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In This Book
This guide includes procedures for setup, operation, and maintenance of the RapidFire System.
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D0004197 July 2021 Revision A.02
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