Agilent 6400 Quick Start Guide

Agilent 6400 Series Triple Quadrupole
LC/MS System

Quick Start Guide

Where to find information 4
Getting Started 6
Step 1. Start the Data Acquisition software 7
Step 2. Prepare the LC modules 13
Step 3. Prepare the Triple Quad instrument 17
Step 4. Set up and run an acquisition method 21
Step 5. Review results with the Qualitative Analysis program 29
Step 6. Set up and run quantitation 29

Use this guide for your first steps with the Agilent 6400 Series Triple Quad LC/MS, and as a
road map for your user information.

What is the Agilent 6400 Series Triple Quad LC/MS system?

The Agilent Triple Quadrupole LC/MS is a liquid chromatograph triple quadrupole
mass spectrometer that performs MS/MS using three sets of parallel rods (in this
case, quadrupole, hexapole, quadrupole). The first quadrupole separates ions into
precursor ions that are fragmented in the hexapole into product ions, which are
separated by the second quadrupole. Often, two or more precursor ions and their
product ions are monitored in sequence in MRM (multiple reaction monitoring)
mode. You can monitor up to 4000 MRM transitions by using Dynamic MRM.

The Agilent 6490 Triple Quadrupole contains the iFunnel Technology which
provides more efficient sampling of ions into the mass spectrometer. The iFunnel
Technology includes the Agilent Jet Stream Technology, shorter desolvation
assembly with Hexabore Capillary, and Dual Offset Ion Funnel.

What’s New in Data Acquisition

The Agilent 6490 and 6460 Triple Quad LC/MS are shipped by default with the
Agilent Jet Stream Technology that utilizes a super-heated sheath gas to collimate
the nebulizer spray which dramatically increases the number of ions that enter the
mass spectrometer.

You can set up an Agilent Triple Quad LC/MS with the Agilent 1100, 1200, 1260 or
1290 Infinity LC modules.

Also, the Agilent Triple Quad LC/MS comes with Agilent MassHunter Workstation
Software that includes three major programs:

• Data Acquisition – From one screen you can tune the mass spectrometer,
control and monitor instrument parameters, set up acquisition methods and
worklists containing multiple samples and monitor real-time run plots.

• Quantitative Analysis – From one screen you can set up a batch of data files and
quantify, evaluate and requantify the results. From this screen you have access
to the Method Editor for setting up and editing the quantitation methods.

• Qualitative Analysis – From one screen you can extract and integrate
chromatograms, subtract background, extract peak spectra, and compare data
from different types of data files.

What’s New in Data Acquisition

The Data Acquisition program for the 6400 Series Triple Quadrupole has many
new features in this revision.

in B.06.00

• The supported models are 6410B, 6420A, 6430A, 6460A, and 6490A.

• Fast Scan is supported on all models.

• The Agilent G7100A Capillary Electrophoresis system is supported.

• The study types Worklist-Only and Worklist Import both support creating a

Quantitative Analysis method.

• The Autotune program has the ability to tune only the unit mass.

• The Autotune program can report m/z below 100 for positive and negative

autotunes.

• You can adjust the EMV separately in the Manual Tune > Detector tab.

• The Tune report contains the instrument serial number and the firmware

revision.

• Triggered MRM databases are supported in Optimizer.

• Additional options for TMRM and compound naming are supported when

searching, filtering and importing to an acquisition method.

2 Agilent 6400 Series Triple Quadrupole LC/MS Quick Start Guide

What’s New in Data Acquisition

• The Source and iFunnel Optimizer program is available to help optimize some
source and ion funnel parameters.

• In Triggered MRM you can select two different triggers for each compound.

• In SIM, MRM, and DMRM methods you can specify a Compound Group.

• You can specify a trigger entrance delay, a trigger time delay and a trigger

window to allow you to acquire triggered scans closer to the apex of the peak.

• Polarity switching is supported for TMRM. Different compounds can have
different polarities.

• If the method is a Triggered MRM method, then you can enable or disable
triggering for all compounds in a method for a specified sample type. For
example, if the Sample Type is Calibration, all of the primary transitions are
acquired, but the secondary transitions are never triggered.

• You can update the threshold value in a method using a Dynamic MRM or
Triggered MRM data file.

• When you switch to Triggered Dynamic MRM, you can automatically set the
default trigger to the transition with the highest m/z (the product ion).

• When you switch from a Triggered Dynamic MRM run to a Dynamic MRM run,
the secondary MRMs are grayed out and are not acquired during the run.

• The Update MRM tool can also fill in the Trigger Window values from a data file
or a Quantitation report. A variety of options are available.

• You can create an MS Indexed data file which supports faster operation in the
Quantitative Analysis program.

• The user interface of the Instrument Status window has been updated.

• The user interface of the DMRM Viewer has been updated.

• The DMRM Viewer supports tMRM methods.

• The DMRM Viewer has added the ability to filter on primaries only or on all

transitions.

• The DMRM Viewer has added Compound and Compound Group navigation.

• In the Study Manager, the Worklist-Only study and the Worklist Import study

include a page to enable quantitation and report generation.

• You can select a post-analysis script that will run after every method in a
worklist; it runs after the Data Analysis portion of the method is complete.

• The 1290 Quaternary Pump (G4204A) is supported.

• The FlexCube (G4227A) is supported in standalone mode.

• UIB 2 (G1390B) is supported.

For a list of updates made to previous versions, refer to the online Help.

Agilent 6400 Series Triple Quadrupole LC/MS Quick Start Guide 3

Where to find information

Help

Where to find information

Help

Press F1 To get more information about a pane, window or dialog box, place the

cursor on the part of the pane, window or dialog box of interest and press F1.

Help menu From the Help menu, access “How-to” help and reference help.

Documents

You can find these manuals delivered with the Triple Quad hardware or software.
You can also find a PDF version on the installation disk, in the Manuals folder.
Many of these manuals are also available on the www.chem.agilent.com

Installation Guide This guide is used to install the hardware and software,

configure the instrument, and verify performance.

Upgrade Guide This guide is used to upgrade your MassHunter Workstation

software from a previous version.

web site.

Maintenance Guide Use this guide to help maintain and troubleshoot your

Agilent Triple Quad LC/MS.

Concepts Guide - The Big Picture Learn the background information to help you

understand operation of the hardware and software.

Data Acquisition for 6400 Series Triple Quad Familiarization Guide Do the

exercises to learn to use the Triple Quad LC/MS hardware and Data Acquisition
program for 6400 Series Triple Quad.

Study Manager Quick Start Guide Use this guide to learn to use the MassHunter

Study Manager software.

MassHunter Optimizer Software Quick Start Guide Use this guide to learn about

the MassHunter Optimizer program. The MassHunter Optimizer program provides
a way to automatically optimize the data acquisition parameters for MRM mode
(multiple-reaction monitoring) on a Triple Quad instrument for each individual
compound analyzed.

4 Agilent 6400 Series Triple Quadrupole LC/MS Quick Start Guide

Training

Where to find information

Training

MassHunter QQQ Compliance Quick Start Guide Use this guide to learn about the

MassHunter QQQ Compliance program.

MassHunter Quant Compliance Quick Start Guide Use this guide to learn about

the MassHunter Quant Compliance program. (This guide is on the Quantitative
Analysis disk.)

Qualitative Analysis Familiarization Guide Do the exercises to learn to use the

Qualitative Analysis program. (This guide is on the Qualitative Analysis disk.)

For information on what is new in the Qualitative Analysis program, refer to the
Qualitative Analysis Familiarization Guide.

Quantitative Analysis Familiarization Guide Do the exercises to learn to use the

Quantitative Analysis program. (This guide is on the Quantitative Analysis disk.)

For information on what is new in the Quantitative Analysis program, refer to the
Quantitative Analysis Familiarization Guide.

Familiarization Guide Use all three familiarization guides to get to know the

software.

Quick Start Guide Use the quick start guides for Study Manager, Optimizer and

Compliance to get to know these programs.

Training Courses Visit www.chem.agilent.com to view a listing of training courses

for the Agilent Triple Quad LC/MS.

Agilent 6400 Series Triple Quadrupole LC/MS Quick Start Guide 5

Getting Started

Start
Here!

Set up, run

and analyze

samples

3 Set up and

run an
acquisition
method

1 Start the

Data
Acquisition
software

2 Prepare the

LC/MS

4 Review results

with
Qualitative
Analysis

5 Set up and run

quantitation

Getting Started

Install the Triple Quadrupole LC/MS hardware and software

Use the Installation Guide to install the hardware and software and verify
performance.

Set up, run and analyze samples

The roadmap below shows you the steps to set up and run a batch of samples from
start to finish. Follow the instructions on the next pages to get started and to learn
where to find the information to help you with each step in this roadmap.

6 Agilent 6400 Series Triple Quadrupole LC/MS Quick Start Guide

Step 1. Start the Data Acquisition software

NOTE

Step 1. Start the Data Acquisition software

The instructions below include the following assumptions:

• The hardware and software are installed.

• The instrument is configured.

Use instructions in the Installation Guide to configure the instrument for the
first time and any time you change the LC configuration.

The LC modules and the Triple Quad MS are turned on, but the LC pump is not
running.

After installation, you see all of the Agilent MassHunter Workstation Software
icons on your Desktop.To start the Data Acquisition program, double-click the
Data Acquisition icon.

The Data Acquisition window appears.

Getting Started

When Data Acquisition opens, the software engines automatically
start. If you need to restart them, right-click the Acq System
Launcher icon in the system tray, and click Start Engines.

If you have recently changed LC modules, remember to configure the
instrument again. See the Installation Guide for instructions.

Agilent 6400 Series Triple Quadrupole LC/MS Quick Start Guide 7

Getting Started

Instrument Status
window

Chromatogram Plot
window

Method Editor
window

Spectrum Plot
window

Actuals
window

The Sample Run and Worklist windows are tabbed here. These three windows are “sharing” this space. You
click the tab to switch to a different window.

Step 1. Start the Data Acquisition software

Windows—where you do most of your work

When you first start the Data Acquisition program, the main window appears. You
do almost all of your work within the eight windows of this main window. These
windows provide the tools to set up acquisition methods, run samples
interactively or automatically, monitor instrument status, monitor runs and tune
the instrument.

Figure 1 Main window of the Data Acquisition software program

8 Agilent 6400 Series Triple Quadrupole LC/MS Quick Start Guide

Getting Started

Step 1. Start the Data Acquisition software

Show/hide the windows You can show one window at a time on the screen or up

to seven windows. You can never hide all of the windows. To show or hide a
window, you click the commands in the View menu. You can also hide a window by
clicking the X icon in the upper right corner of the window.

When you click a window, the title of the active window changes to a different
color. Press F1 to obtain help on the active window. You can also drag a window
border to resize the window. If you double-click the title of the window, the
window “floats” outside of the main window. You can double-click the title bar
again to “dock” the window. You can also float and dock the window by
right-clicking the title of the window and clicking Floating.

Instrument Status window With this window you view the status of each device

configured with the instrument- Error, Not ready, Pre-run, Post-r un, Running,
Injecting, Idle, Offline, or Standby. You also set non-method control and

configuration parameters for the LC devices and the MS instrument.

This window displays each device's current status both as text and by its
color-coding:

Color Coding in the Instrument Status Window

Color Status

Red Error

Yellow Not ready

Purple Pre-run, post-run, Waiting

Blue Running, injecting

Green Idle

Dark gray Offline

Light gray Standby (for example, lamps off)

Agilent 6400 Series Triple Quadrupole LC/MS Quick Start Guide 9