acquity uplc sample manager - fixed loop operator’s ... · valve a three-position, rotary-shear...

ACQUITY UPLCSample Manager - Fixed Loop Operator’s Overview and Maintenance Information

Revision A

Copyright © Waters Corporation 2010–2011All rights reserved

ii

Copyright notice

© 2010–2011 WATERS CORPORATION. PRINTED IN THE UNITED STATES OF AMERICA AND IN IRELAND. ALL RIGHTS RESERVED. THIS DOCUMENT OR PARTS THEREOF MAY NOT BE REPRODUCED IN ANY FORM WITHOUT THE WRITTEN PERMISSION OF THE PUBLISHER.

The information in this document is subject to change without notice and should not be construed as a commitment by Waters Corporation. Waters Corporation assumes no responsibility for any errors that may appear in this document. This document is believed to be complete and accurate at the time of publication. In no event shall Waters Corporation be liable for incidental or consequential damages in connection with, or arising from, its use.

Table of Contents

Copyright notice ........................................................................................................ ii

Overview .................................................................................................................. 1 Location of SM-FL in the ACQUITY UPLC system .......................................... 1 Flow path through the sample management system......................................... 2 SM-FL major components ................................................................................... 3 Functional systems .............................................................................................. 6

Preparing for operation ........................................................................................... 9 Installing the optional leak sensor ..................................................................... 9 Installing the waste tubing ............................................................................... 11 Priming the SM-FL............................................................................................ 13

Using the SM-FL ................................................................................................... 15 Interface requirements ...................................................................................... 15 Installation recommendations for fittings........................................................ 16 Selecting weak wash and strong wash solvents............................................... 20 Washing the SM-FL needle ............................................................................... 21 Loading sample plates ....................................................................................... 23 Sample chamber considerations ....................................................................... 27 Choosing needles and sample loops .................................................................. 27 Choosing the sample syringe............................................................................. 30 Choosing the sample syringe draw rate ........................................................... 31 Choosing the needle height setting................................................................... 32 Recovering maximum sample from vials.......................................................... 33 Revising a plate type ......................................................................................... 34 Air gaps .............................................................................................................. 35 Selecting the optimum sample injection mode................................................. 36 Load ahead option.............................................................................................. 40 Loop offline option ............................................................................................. 41 Diagnostic tests.................................................................................................. 44 Resolving leak sensor errors ............................................................................. 45

Table of Contents iii

Maintaining the SM-FL ........................................................................................ 49 Contacting Waters technical service................................................................. 49 Maintenance schedule ....................................................................................... 50 Maintenance considerations.............................................................................. 51 Configuring maintenance warnings ................................................................. 52 Replacing the leak sensor.................................................................................. 52 Replacing the sample needle ............................................................................. 55 Calibrating the needle’s z axis .......................................................................... 63 Replacing the sample loop................................................................................. 64 Replacing the sample syringe ........................................................................... 67 Replacing the wash syringes ............................................................................. 70 Replacing the injection valve cartridge ............................................................ 74 Cleaning the instrument’s exterior................................................................... 76

iv Table of Contents

Overview

You can submit samples for analysis on the ACQUITY UPLC system by loading microtiter plates or vials onto the rotary sample tray of the sample manager-fixed loop (SM-FL). Using a fixed loop, the sample manager injects the samples it draws from the plates and vials onto a chromatographic column. Optional sample loops make possible a range of injection volumes.

Location of SM-FL in the ACQUITY UPLC system

The following diagram shows the location of the SM-FL in the ACQUITY UPLC system.

Bottle tray

Detector

Column heater

Sample Manager - Fixed Loop

Solvent manager

Overview 1

Tips:

• Use care when stacking or moving the SM-FL. Ensure the drip tray does not collide with any surface.

• Use care when installing and removing reusable fittings.

See also: Column Compartments Operator's Overview and Maintenance Information on the ACQUITY UPLC System Documentation CD to avoid potential leaks or carryover.

Flow path through the sample management system

The following diagram shows how the SM-FL functions as part of the ACQUITY UPLC system.

Drainage from column heater

SampleDegassed solvent from solvent manager

Inject valve

Mobile phase from solvent manager

To column

Secondary drainage to solvent

manager

Secondary drainage to

solvent manager

Primary drainage to solvent manager

2

SM-FL major components

The following diagrams show the SM-FL’s major components.

Front view with doors closed:

On/off switch

Power LED

Run LED

Sample compartment doorFluidics compartment door

Overview 3

Front view, with doors open:

Plate selector switch

Sample syringe

Location of sample manager leak sensor

Injection valve

Sample tray

Chamber temperature sensor

Access panel

Sample syringe valve

Location of column heater leak sensor

Location of volume detection device

4

Sample compartment components visible with access panel removed:

SM-FL components:

Component Description

Access panel Removable panel (requires a TORX® driver) that allows access to sample compartment components such as the seal assembly and needle carriage.

Compartment lighting

LED that illuminates the sample compartment. The compartment light automatically turns on when the SM-FL’s sample compartment door is opened and turns off when the door is closed. The light can also be turned off via the console to accommodate light-sensitive samples.

Injection valve A two-position, six-port injection valve.

Volume detection device

A sensor that detects air, versus liquid, in a clear tube, determining the volume of the installed sample needle and sample loop.

Location of sample needle

Sample needle carriage

Compartment lighting

Location of column heater leak sensor cable connector

Location of sample manager leak sensor cable connector

Overview 5

Functional systems

RΘ positioning mechanism

The RΘ (R-theta) positioning mechanism’s two axes control the orientation of the sample plates within the sample compartment and the relative position of the sample needle carriage. The theta-rotary axis is a belt-driven shaft that rotates a pair of sample plates 360° from a reference point. The R-linear axis is the axis along which the sample needle carriage is oriented. The carriage runs from the rear-left corner to the front-right corner of the sample compartment.

Injection system

The injection flow path includes the assemblies required to aspirate a sample and deliver it to the column. The process involves the needle, sample loop, sample syringe and syringe valve, and injection valve.

Leak sensor Continuously monitors the SM-FL for leaks and stops the system flow when its optical sensor detects about 1.5 mL of accumulated, leaked liquid in its surrounding reservoir.

Plate selector switch Toggle switch used to select either plate position 1 or plate position 2.

Sample syringe Draws sample into the sample needle.


A three-position, rotary-shear valve.

Sample needle Extracts sample from vials.

Sample needle carriage

Positions the sample needle in the sample compartment.

Sample tray Secures the sample plates or vial holders in place.

Chamber temperature sensor

Located on the upper guide rail toward the back of the sample chamber, to monitor sample environment.

SM-FL components: (Continued)

Component Description

6

Wash system

The wash system uses a wash pump, wash solenoid, and weak-wash and strong-wash syringes to rinse areas that are contacted by the sample.

The system uses two wash solvents:

• Strong needle-wash solvent, for flushing injected components from areas that are contacted by the sample. Strong needle-wash solvent is never injected along with the sample.

• Weak needle-wash solvent, for transporting sample to the sample needle and eliminating strong needle-wash solvent. Weak needle-wash solvent is injected with the sample in partial-loop, pressure-assist mode.

The wash sequence consists of five primary operations:

1. The needle moves to the drain position and empties any excess sample into the drain port. The volume of wash solvent consumed during this flush depends on the size of the sample, air gap, and loop.

2. The needle remains in the drain position for rinsing with strong needle-wash solvent:

• If the specified volume of strong needle-wash solvent is fewer than three times the needle volume (about 150 µL), the entire wash volume is dispensed at the drain position.

• If the specified volume of strong wash exceeds three times the needle volume, the remainder (wash volume minus three times the needle volume) is dispensed at the combined injection and wash port.

As the needle dispenses strong wash solvent, an equal amount of weak needle-wash solvent forces upward through the bottom of the combined injection and wash port.

3. The weak wash flushes the needle, removing from it any remaining traces of strong wash solvent.

4. The wash syringes are replenished with degassed wash solvent from the solvent reservoirs.

5. The needle is briefly rinsed at the drain when the sample loop returns to the load position, which happens at the start of a partial loop injection with pressure assist or a full loop mode injection. The brief rinsing ensures the loop, which is at system pressure rather than atmospheric

Overview 7

pressure, is clear of any residual sample that could be expelled when the pressure is released.

Flow path diagram:

8

Thermal system

The thermal system maintains the set temperature in the sample compartment.

Tips:

• You do not need to defrost the sample compartment.

• The SM-FL’s fans stop circulating air whenever the sample compartment door is open.

• The sample tray rotates slowly when the system is idle, to help maintain a uniform temperature across the plates.

Preparing for operation

Note: The system is shipped with a 10-μL PEEK™ needle. If you are not using the default ACQUITY UPLC system configuration, which uses this needle, see page 49.

Before you prepare the SM-FL for operation, prepare the solvent manager.

Tip: For instructions on preparing the solvent manager, see ACQUITY UPLC Binary Solvent Manager Operator’s Overview and Maintenance Information.

Preparation of the SM-FL involves these steps:

• Installing the leak sensor

• Installing the waste tubing

• Priming the SM-FL

Installing the optional leak sensor

Warning: To avoid the harmful effects of personal contact with solvents, including inhalation, observe Good Laboratory Practice when you handle them. See the Material Safety Data Sheets for the solvents you use.

Warning: To avoid personal contamination with biologically hazardous or toxic materials, wear clean, chemical-resistant, powder-free gloves when performing this procedure.

Preparing for operation 9

Required materials

• Gloves: clean, powder-free, chemical-resistant

• Leak sensor

To install the leak sensor:

1. Power-off the SM-FL.

2. Open the fluidics compartment door.

3. Carefully unpack the new leak sensor.

4. Align the leak sensor’s T-bar with the slot in the side of the leak sensor reservoir, and slide the leak sensor into place.

5. Plug the leak sensor connector into the front of the instrument.

Caution: To avoid damaging electrical parts, never disconnect an electrical assembly while power is applied to an instrument. To completely interrupt power, set the power switch to Off, and then unplug the power cord from the AC source. Wait 10 seconds thereafter before you disconnect an assembly.

TP02892

T-bar

Leak sensor installed in reservoir

Slot in leak sensor reservoir

10

6. Power-on the SM-FL.

7. In the ACQUITY UPLC Console, select Sample Manager FL from the system tree.

8. In the SM-FL information window, click Control > Reset SM, to reset the SM-FL.

Installing the waste tubing

Required material


• ACQUITY UPLC I-Class System Accessories kit

To install the waste tubing:

1. Attach the corrugated tubing, provided in the system accessories kit, to the process waste port (found on the lower drip tray of the sample manager).

Caution: To prevent contamination to system components, wear clean, chemical-resistant, powder-free gloves when installing or removing the waste tubing.


2. Route the tubing through the pass-through on the upper drip tray of the solvent manager.

3. Slide the adapter onto the end of the corrugated Teflon® tubing.

4. Connect the adapter to the front boss fitting on the lower drip tray of the solvent manager.

Tip: For instructions on routing the solvent manager waste and vent lines, see the Binary Solvent Manager Operator’s Overview and Maintenance Information.

Corrugated Teflon tubing

Process waste port

Pass-through on upper drip tray of the solvent manager

Front boss fitting on lower drip tray of the solvent manager

12

Priming the SM-FL

The priming process fills the sample needle with solvent, flushes new solvent through the injector lines, and/or purges air from the lines. You prime the system to accomplish these tasks:

• Prepare a new SM-FL for operation

• Prepare a SM-FL for operation after it has been idle for an extended period

• Change the solvent in the syringes

• Remove bubbles from the lines

Ensure that the priming solvent is correctly composed and that it is high in quality and miscible with any other solvents used in your system. Use filters in all solvent reservoirs, and ensure the volumes of solvents are sufficient for priming.

To prime the sample syringe:

1. In the ACQUITY UPLC Console, select Sample Manager from the system tree.


Sample Manager information window:

2. Click Control > Prime Syringes.

Alternative: Right-click in the SM-FL control panel, in the data application, and then click Prime.

3. In the Prime Syringes dialog box, select Sample syringe and both wash syringes.

Tip: If you want only to remove air bubbles from the sample syringe, but do not want to prime the wash syringes, select Sample syringe only. However, do not select this option routinely. Priming all components at the same time is good practice.

4. Type the number of primes in the Number of cycles text box.

Default: 1

Recommendation: Specify 5 to 7 primes when you are changing solvents.

14

5. Click OK to start priming. When the system status is “Idle,” priming is finished.

Tip: Each prime takes approximately 2 to 4 minutes.

Using the SM-FL

Before running samples,

• Examine the injection valve, sample syringe, and all fittings for leaks. Tighten fittings as needed.

• Ensure both the sample compartment door and the fluidics compartment door are closed.

Interface requirements

Ventilation

Allow at least 15.2 cm clearance at the rear and at least 1.3 cm clearance on the right-hand side of the SM-FL for ventilation.

Drainage system

• Allow clearance for fluid lines to pass along the right-hand side of the instrument (twelve 0.125-inch OD tubes or four 1/6-inch OD tubes, for example).

• Ensure that the SM-FL can accept drainage from the column heater and provide a path to waste.

Caution: Do not abort the sample manager priming sequence. Doing so can leave strong solvent in the sample needle, which can adversely affect chromatography.

Using the SM-FL 15

Installation recommendations for fittings

Most tubing connections use gold-plated compression screws and two-piece ferrules. See the diagram below for assembly orientation.

Gold-plated compression fitting

The sample needle uses a PEEK compression screw and two-piece ferrules. The compression screw differs slightly, depending on the sample needle’s outside diameter. See the diagram below for assembly orientation.

PEEK sample needle fitting

The sample needle fitting is sometimes preset on the needle. Where it is not preset, assemble the fitting as shown above, ensuring that the thin wall of the

Warning: To avoid personal contamination with biologically hazardous or toxic materials, wear clean, chemical-resistant, powder-free gloves when reinstalling fittings.

Caution: When installing or removing a column, be sure that you turn the column itself and not the fitting (compression screw and ferrule). Otherwise, the active pre-heater tubing can turn with the fitting, potentially damaging the active pre-heater assembly.

TubingCompression screwFerrule with locking ring

TubingCompression screw

Ferrule

Locking ring (thin edge toward ferrule)

16

locking ring faces the flat-bottom ferrule. Doing so ensures the ferrule is correctly compressed onto the needle tubing.

Recommendations:

• To prevent band spreading, ensure the tubing bottoms in its fitting hole before you tighten the compression screw.

• For easier accessibility, use long compression screws to attach tubes to the injector and vent valve.

• Perform the sample syringe leak test whenever you replace or loosen fittings during maintenance (see the ACQUITY UPLC online Help).

• Whenever you loosen fittings during maintenance, examine them for cracks, stripped threads, and deformations.

• Do not reuse stainless steel fittings more than six times.

Required material

Gloves: clean, powder-free, chemical-resistant

When tightening system fittings, consult the following table.

Installation recommendations for ACQUITY UPLC fittings:

Fitting Recommended tightening

Fir

st u

se o

rre

-in

stal

led 1/4-28 flangeless with ferrule Finger-tight

Fir

st u

se

1/4-28 flangeless with 2-piece ferrule Finger-tight

Re-

inst

alle

d 1/4-28 flangeless with 2-piece ferrule Finger-tight

Using the SM-FL 17

Fir

st u

se o

rre

-in

stal

led 10-32 one-piece PEEK Finger-tight

Fir

st u

se

Stainless steel (gold-plated) with 2-piece stainless steel ferrule

Finger-tight, plus 3/4-turn using wrench

Re-

inst

alle

d

Stainless steel (gold-plated) with 2-piece stainless steel ferrule (re-installed)

Finger-tight, plus up to 1/6-turn using wrench

Fir

st u

se w

ith

u

nas

sem

bled

fer

rule

PEEK fitting with flat-bottom ferrule Finger-tighten until first resistance is felt, then tighten an additional 1/4-turn using fingers or wrench.

Installation recommendations for ACQUITY UPLC fittings: (Continued)


3/4-turn

1/6-turn

1/4-turn

18

Fir

st u

se w

ith

pr

e-se

t fe

rru

lePEEK fitting with flat-bottom ferrule Finger-tighten until first

resistance is felt, then tighten an additional 1/4-turn using fingers or wrench.

Re-

inst

alle

d

PEEK fitting with flat-bottom ferrule Finger-tighten until first resistance is felt, then tighten an additional 1/4-turn using fingers or wrench.

Installation recommendations for ACQUITY UPLC fittings: (Continued)


1/4-turn

1/4-turn

Using the SM-FL 19

Selecting weak wash and strong wash solvents

For best performance, follow these guidelines when selecting wash solvents. Otherwise performance can be reduced, specifically Area/Height RSD and Linearity. Careful selection of wash solvents can also reduce carryover. The guidelines do not prohibit all other solvent combinations. Other combinations can be run with lower performance expectations or by manipulating default injection parameters.

Use a weak wash solvent based on the sample and mobile phase chemistries of your application, making sure all solutions/buffers are miscible and soluble.

Recommendation: For buffered aqueous, reversed-phase chromatographic conditions and MS applications, use a weak wash solvent of 100% water or 0% to 25% methanol or acetonitrile and a strong wash solvent of 50% to 100% methanol or acetonitrile. High sample concentrations can require other weak wash solvents. If your separation permits, Waters recommends adding a small amount of an organic solvent (~10%) to prevent microbial growth.

See the Solvent Considerations appendix in the ACQUITY UPLC System Guide for further information about solvents.

Tips:

• For best performance, the weak wash solvent must be similar or identical to your isocratic or initial gradient solvent conditions, excluding buffers. Do not use salt buffers in wash solvents.

• Choose a strong solvent at least equivalent to your final gradient conditions. A good starting point is a mixture of 25% water/25% acetonitrile/25% methanol/25% isopropanol.

Caution: To avoid damage to the solenoid valve seats and seals in the solvent path, do not use a nonvolatile buffer as the weak wash or strong wash solvent.

20

Washing the SM-FL needle

Washing the needle is an optional procedure that flushes strong and/or weak wash solvent through the needle and injection port. Washing the needle removes contaminants from the inside and outside of the needle, the external piercing needle, and the injection port. You can also perform a needle wash to ascertain proper flow through the waste tubing and to confirm that the needle wash system is primed and properly operating.

Wash solvent effects:

Property Effect

Organic species As a general principle, strong and weak solvents must include the same organic species. Note that this is not always practicable. You can, however, use a 100% organic strong wash solvent.

pH Adjust the pH of the strong and weak solvents for best peak shape and carryover performance.

Concentration of strong solvent

The strong solvent offers different selectivity than the weak wash solvent. The strong solvent must be no stronger than the concentration needed to reduce carryover to an acceptable level.

Sample diluent The weak wash solvent will contact the sample, so match these as closely as possible. To offset adverse effects on peak shape caused by the matrix’s composition, adjust the weak wash composition, especially when using the instrument in partial loop mode.

Wash volume ratio (weak to strong)

Within a method, use a 3:1 ratio, weak wash to strong–sufficient to ensure the weak wash flushes the strong from the needle and sample loop before sampling.When using a viscous strong solvent, use a 6:1 to 10:1 ratio, weak wash to strong.

Cycle times Higher viscosity wash solvents lengthen wash cycles.

Using the SM-FL 21

Tip: Priming the system washes the sample needle, so whenever you prime the system, you can omit this procedure.

To wash the SM-FL needle:


2. In the SM-FL information window, click Control > Wash Needle.

Alternative: Right-click the SM-FL control panel in the data application, and then click Wash Needle.

3. In the Strong Wash box, specify the volume for the strong wash solvent. Or, to omit strong wash solvent, enter 0 in the Strong Wash box, or leave it blank.

Range: 0.0 through 99,999 μL

Default: 0.0 μL

Recommendation: 100 through 500 μL

Tip: Using both a weak and strong wash solvent increases the wash time and solvent consumption because the system must be fully cleansed of the strong solvent before starting the next injection.

4. In the Weak Wash box, specify the volume for the weak wash solvent.

Range: 1.0 through 99,999 μL

Default: 200.0 μL without strong wash or 500 μL with strong wash

Recommendation: 200 through 500 μL or three times the strong wash volume.

5. Click OK.

Result: The needle wash begins. When it ends, the status returns to idle.

Caution: To avoid strong wash solvent contacting the sample and contaminating it, use a sufficient quantity of weak wash solvent.

22

To stop a needle wash routine before it finishes:

In the SM-FL information window, click Control > Reset SM.

Alternative: Right-click the SM-FL control panel in the data application, and then click Reset SM.

Loading sample plates

The SM-FL is compatible with the ANSI standard well-plates, vial-trays, vials, and cap-mats/seals that are approved for use with the ACQUITY UPLC system. The SM-FL holds two ANSI/SBS plates that you load through the sample compartment door.

Requirement: The plates you use must meet ANSI/SBS standards.

Tip: Vial positions V1 through V4, located on the right-hand and left-hand sides of the sample tray, accommodate 4-mL vials. Contact Waters for inserts that allow you to use 2-mL vials in these positions.

Observing vial and plate recommendations

Waters recommends that you observe these usage guidelines for sample vials and plates in the SM-FL:

• Vials

– Use only Waters-certified vials.

– Ensure that vial holders conform to ANSI/SBS standards.

• Plates

– Use only 1860024XX-series plates and cap mats in the SM-FL.

– When selecting a new plate supplier, especially for 384-well plates, measure the plate size to ensure compatibility with Waters’ specifications for the SM-FL.

– To avoid warping plates, do not centrifuge them.

– Be aware that plates containing samples with high concentrations of organic solvent can give inconsistent results at room temperature.

Caution: Do not abort the sample needle wash sequence. Doing so can leave strong solvent in the sample needle, which can adversely affect chromatography.

Using the SM-FL 23

• Covers

– Use foil covers on vial plates whenever possible.

– Use pre-slit cap-mats/seals and vial caps. Use of non-pre-slit cap mats and vial caps can cause clogging in the wash lines.

– To prevent sample spillage or needle damage, use only Waters-approved covers on the sample vials.

For more information about plates and vials, see Using Plates and Vials with ACQUITY UPLC Systems, part number 715002434.

Required material

Safety glasses

To load a sample plate:

1. Open the sample compartment door.

2. Press the plate selector switch on the top, center of the door frame to select plate position 1 or 2.

Exception: If the SM-FL is accessing the sample tray when you select a new plate position, the selector switch does not operate and the instrument beeps once. The selector switch operates again after the SM-FL no longer accesses the sample tray.

Tip: Press the plate selector switch twice to toggle between loading a sample plate and loading positions that accept 4-mL vials.

Warning: To avoid eye injury, wear safety glasses when loading sample plates.

24

3. Pull the sample tray out.

4. Load the plate onto the tray so that well position A,1 is at the rear, left-hand corner, and the forward edge of the plate is behind the spring inside the front of the carrier.

Tip: “A” represents the row number, “1” represents the vial position.

Sample tray pulled out

Position for a 4-mL vial (V1 through V4)

TP03232

Sample plate

A,1 well position

Using the SM-FL 25

Sample plate vial positions:

5. Slide the tray in until it clicks into place.

6. Close the sample compartment door.

Caution: To avoid damaging the sample needle, the sample plates must be positioned correctly and the sample tray must be fully engaged.

1 2 3 4 5 6 7 8

A

B

C

D

E

F

Sample tray handle

26

Sample chamber considerations

Choosing needles and sample loops

Sample needles

Waters offers multiple needle sizes so you can choose the best option for your injection volume and sample viscosity. Waters recommends metal needles for samples known to be attracted to hydrophobic polymers and also when you use hexane and tetrahydrofuran.

Tip: Using a smaller needle increases cycle times.

Warning: To avoid puncture wounds, keep hands or loose clothing clear of the needle assembly mechanism while it is moving. Note that the SM-FL beeps three times whenever the sample compartment door is open, and the needle assembly mechanism is about to move.

The following sample needles are available for the SM-FL:

Needle size (μL)

Material

10 PEEK (17-inch L × 0.007-inch ID) – Default

10 PEEKsil™

10 FEP with stainless steel tip

10 stainless steel

20 PEEK (17-inch L × 0.010-inch ID)

20 PEEKsil™

20 FEP with stainless steel tip

20 stainless steel

Using the SM-FL 27

Follow these guidelines to select the most appropriate needle material for your application:

Sample loops

The sample loop holds the sample for injection. You install the sample loop between port 1 and port 4 on the injection valve.

Sample loop volume is calculated based on loop length and an assumed internal diameter (ID), which can vary by as much as 0.001 inch in stainless steel tubing. This variation in internal diameter across the length of the loop tubing can result in significant variation in the actual loop volume.

Needle material Recommended for

PEEK General use

PEEKsil Use with hydrophobic compounds

Stainless steel Use with plates and capmats

FEP with stainless steel tip Use with plates and capmats

The following sample loops are available for the SM-FL:

Loop size (μL)

Minimum volume (μL) Maximum volume (μL)

1 0.78 1.31

2 1.53 2.55

5 4.31 5.78

10 9.01 11.10

20 16.76 23.53

50 40.45 50.55

100 86.47 100.66

250 217.25 250.78

28

The table below illustrates the impact of variation in internal diameter for a 5-µL sample loop constructed of 0.009-inch (nominal) ID tubing.

To observe the volume variation in the installed sample loop:

1. In the ACQUITY UPLC Console, click the Sample Manager view.

2. Select Maintain > Characterize > Needle and loop volumes.

3. Compare the measured and the nominal loop volumes.

Modifying needle and sample loop configuration parameters

To configure the system for a needle or sample loop size that differs from the one currently fitted:


2. Select Configure > Volumes.

3. In the Volume Configuration dialog box, select the appropriate needle or sample loop size from the list, and then click OK.

Needle and sample loop characterization

To ensure correct positioning of the sample in the sample loop, the sample manager “characterizes” the installed loop and needle. In doing so, it determines their actual volume, minimizing error associated with intrinsic variations in the volume of those components. The characterization time varies depending on the size of the installed loop and needle.

To ensure accurate injection volumes, the sample manager slows the aspiration rate to 10 µL/min during characterization. It then determines the volume of the loop by locating the position of the aspirated air bubble twice, once with the loop offline and once with the loop online. For a 30-µL needle,

Internal diameter (inch)

Volume per inch length (μL)

Nominal to actual volume ratio

Volume (μL)

0.009 1.04 1 5

0.010 1.28 1.23 6.2

0.0105 1.42 1.37 6.9

Using the SM-FL 29

the air gap must displace about 40 µL of fluid to reach the volume detection device. The displacement process requires approximately four minutes for each mode, eight minutes in total.

Because variations in the tolerance of the sample loop's internal diameter become more pronounced as the internal diameter of the loop tubing decreases, smaller loops can require more characterization time than larger loops. Solvent travels more slowly through the tubing of smaller loops, slowing the characterization process.

UV-absorbing mobile phases, such as those containing TFA, can impede detection of the aspirated air gap, producing unsatisfactory characterization results. If the result of the characterization process is unsatisfactory, the sample manager repeats the process.

Choosing the sample syringe

The following sample syringe sizes are available to use in the SM-FL:

• 50 μL

• 100 μL – Default

• 250 μL

• 500 μL

Choose a syringe size that allows you to draw your desired total sample volume into the sample needle. Waters recommends using a sample syringe volume that is at least two times your sample volume.

Modifying sample syringe configuration parameters

To configure the system for a syringe size that differs from the one currently fitted:


2. Select Configure > Volumes.

3. In the Volume Configuration dialog box, select the appropriate sample syringe size from the list, and then click OK.

30

Choosing the sample syringe draw rate

The ideal syringe draw rate depends on sample volume, sample viscosity, and desired cycle time. The default draw rate varies depending on the volume of the sample loop and the needle installed in the device.

You can also specify the draw rate, if desired. The following tables list the maximum draw rates for common mobile phases.

Default draw rates:

Loop volume (µL)Draw rate (µL/minute)

10-µL needle 20-µL needle

1 10 11

2 24 33

5 45 90

10 (default) 60 185

20 60 185

50 60 185

100 60 185

250 60 185

Maximum draw rates for 10-µL needle:

Solvent

Draw rate (µL/minute)

1 µL loop 2 µL loop 5 µL loop10 µLor larger loop

50% MeOH/ 50% H2O

10 24 45 60

100% H2O 30 45 85 120

100% ACN 80 120 220 310

100% DMSO 13 20 37 52

Using the SM-FL 31

Choosing the needle height setting

The default needle penetration depth setting of 2.0 mm prevents you from reaching the bottom of the vial.

You can change the default needle placement (needle height) setting in the software in two places: the Dilution tab of the Sample Manager instrument method editor and the Advanced Settings dialog box. Set consistent values in both places.

To aspirate more sample from the vial, decrease the value so that the needle tip is closer to the bottom of the vial.

Maximum draw rates for 20-µL needle:

Solvent

Draw rate (µL/minute)

1 µL loop 2 µL loop 5 µL loop10 µLor larger loop

50% MeOH/ 50% H2O

11 33 90 185

100% H2O 37 60 170 400

100% ACN 100 160 450 500

100% DMSO 16 27 75 185

Caution: To avoid damaging the needle, follow the guidelines in this section, and use the appropriate needle-height setting for your sample plates or vials.

32

Recovering maximum sample from vials

The current ANSI plate (48 vials) definition for the 2-mL Maximum Recovery Vials can leave some sample in the vial. If you want to recover the maximum amount of sample, change the needle placement setting.

Tip: To modify needle placement for vials, click Instrument Method Editor > ACQ-FL > General tab > Advanced, and change the “Needle Placement (from bottom)” value.

Vial type

Minimum needle placement (mm)

Description Part number

Waters Supplied Total Recovery Vial

0.7 Screw Cap 12 × 32 mm Clear Total Rec pre-slit PTFE/Silicone Septa

186000385C

Waters Supplied Max. Recovery Vial

2.1 Screw Cap 12 × 32 mm Clear Max Rec pre-slit PTFE/Silicone Septa

186000327C

Screw Cap 12 × 32 mm Amber Max Rec pre-slit PTFE/Silicone Septa

186003886C

Vial depth

Sample needle

2 mm offset

Using the SM-FL 33

See also: Waters Sample Vials and Accessories brochure on www.waters.com.

Revising a plate type

The ANSI-48Tube0.65mLHolder plate type is defined in Empower software so that the needle does not use sample near the bottom of the vial, greatly increasing the residual volume. The plate type contains 0.65 mL tubes.

To create a new plate type and correct the depth value:

1. In Empower software, open the Configure System window.

2. From the Empower Configuration tree, select Plate Types.

3. Select the plate type “ANSI-48Tube0.65mLHolder”.

4. From the menu, select File > Properties.

5. Under Plate Type, type a suitable name for the plate.

6. Change the Depth parameter to 28.5.

7. Click OK.

Result: A copy of the plate definition is created with a new name and revised needle depth.

Waters Supplied Flat Bottom Vial

0.1 Screw Cap 12 × 32 mm Clear with pre-slit PTFE/Silicone Septa

186000307C

Screw Cap 12 × 32 mm Amber pre-slit PTFE/Silicone Septa

186000847C

Screw Cap 12 × 32 mm 750 µL PP pre-slit PTFE/Silicone Septa

186002636

Screw Cap 12 × 32 mm 300 µL PP pre-slit PTFE/Silicone Septa

186002639

Vial type

Minimum needle placement (mm)

Description Part number

34

Air gaps

If air gaps are required for your chromatography, you can specify an air-gap volume in the instrument method editor.

The default air gap volume is dependent on the volume of the installed needle and the selected injection mode.

To specify an air-gap volume:

1. In the instrument method editor, click the ACQ-FL tab and then the General tab.

2. Click Advanced.

3. In the Advanced Settings dialog box, mark the box for the air-gap option.

4. Specify volumes for your pre-aspirate and post-aspirate air gaps, and then click OK.

Default air gap volume:

Needle volume

Default air gap volume (µL)

Partial loop needle overfill mode

Partial loop mode Full loop mode

10 µL 1 2 2

20 µL 1 4 4

Using the SM-FL 35

Selecting the optimum sample injection mode

The sample manager supports three injection modes:

• Full loop

• Partial loop

• Partial loop needle overfill (PLNO)

The injection process differs for each mode and thus, each mode has a different accuracy, visualized as a difference in peak area. Because the injection process differs, each mode delivers different amounts, albeit reproducibly and with linearity. The responses cannot be compared or plotted on the same calibration curve.

Each injection mode has different advantages and can be used to improve performance, depending upon the individual goal of the analysis.

Injection mode comparison:

Injection mode Advantages Disadvantages

Full loop • Best precision.• Lowest dispersion.

• Increased sample consumption.

• Multiple sample loops are required for multiple injection volumes.

• Overfilling the loop lengthens the cycle time.

Partial loop • Lowest sample consumption.

• Good linearity and precision.

• Reduces cycle time slightly, compared to partial loop needle overfill mode.

Weak wash solvent, mobile phase, and sample are co-injected, and thus sample peak recoveries are method dependant.

36

Full loop mode

The full loop injection mode (recommended when using 1.0-mm I.D. columns) permits the least peak dispersion and delivers the highest precision of all the modes, particularly when you use it with smaller loops. When used with a 1-µL sample loop or a 2-µL sample loop, it is the best mode for plate-count studies.

Because loop volume typically varies slightly, even for loops of the same nominal loop volume, area values can vary depending on which system an analysis is run. However, accuracy is determined by the tubing's internal diameter tolerance, not the injector performance. See page 28 for more information about loop volume variation.

Partial loop needle overfill

• Good linearity, precision and accuracy.

• Yields consistent sample accuracy and peak areas from system to system and between sample loops.

• Offers lower dispersion than partial loop pressure assist mode injections.

• Sample and its diluent are exposed only to the weak wash and initial mobile phase.

• Only mobile phase and sample are injected.

• The sample is positioned at end of sample loop closest to column inlet.

• Increased sample consumption.Note: Under default conditions, uses 7 µL additional sample per injection.

Injection mode comparison: (Continued)

Injection mode Advantages Disadvantages

Using the SM-FL 37

Partial loop mode

In partial loop mode, sample is drawn, placed between pressurized air gaps and then transferred to the center of the sample loop. Bracketing the sample volume with air gaps (bubbles) reduces peak dispersion resulting from laminar flow. To further reduce dispersion, and to retain the maximum amount of sample in loop, the sample is positioned with its leading edge at the end of the sample loop closest to the column.

Use partial loop mode when analysis time takes precedence over any other concern, when the sample volume is very limited, or when the injection volume is large.

In partial loop mode, weak wash solvent and sample are co-injected, so sample peak recoveries depend on the method used and on these conditions:

• Sample viscosity and the syringe draw rate

• The ratio of the sample volume to the loop volume. For best recovery, keep the ratio between 10% and 50% of the nominal loop volume.

• The ability of the sample manager to accurately characterize the volume of the sample needle, sample loop, and connecting tubing. To improve recovery, characterize the system using the solvents in the current method.

Because the sample is co-injected with the weak wash solvent, ensure that the weak wash solvent does not interfere with the chromatography.


In partial loop needle overfill mode, the sample needle draws extra sample to rinse the needle, ensuring that the volume placed into the sample loop is homogeneous. Partial loop needle overfill mode is the best general-purpose injection mode. It confers excellent linearity performance, and provides superior partial loop accuracy, precision, and linearity for a wide range of samples, including strong and weak acids and bases, hydrophilic, and hydrophobic compounds.

The portion of the sample injected is the least diluted part of the sample volume. Only the initial mobile phase and sample are placed in the sample loop. Therefore, both the strong and weak wash can be used to ensure minimum carryover.

This mode provides better recovery than partial loop pressure assist mode does. The loop is pre-rinsed, replacing the weak wash with extra sample. The

38

result is a less diluted sample than in partial loop pressure assist mode, where the sample slug is introduced by displacing the weak wash, diluting the sample.

Partial loop needle overfill mode provides optimum performance when injection volumes are maintained within a range of 10% to 75% of the actual loop volume. The generally accepted practice for partial loop injection linearity restricts the injection volume selected to ≤ 50% nominal loop volume. Nevertheless, you can increase the usable loop volume to ≤ 75% of the actual loop volume by selecting the needle overfill technique. Doing so is based on achieving an injection-to-injection variability of ≤ 1% across the specified volume range. In addition, the correlation between specified injection volume and peak area is R2 > 0.999.

The following table provides the minimum and maximum injection volumes for each of the commonly used loops.

Selecting the injection mode and loop volume:

Loop volume

Sample injection volume range (µL)


Partial loop mode Full loop mode

1 µL 0.1 to .8 Not recommended 1

2 µL 0.2 to 1.5 Not recommended 2

5 µL 0.5 to 3.8 Not recommended 5

10 µL (default) 1.0 to 7.5 1.0 to 5.0 10

20 µL 2.0 to 15.0 2.0 to 10.0 20

50 µLa

a. You must install a 250- or 500-μL sample syringe in the sample manager when using this size sample loop.

5.0 to 37.0 5.0 to 25.0 50

100 µLa 10 to 75.0 10 to 50 100

250 µLa 25 to 187.5 25 to 125 250

Using the SM-FL 39

Load ahead option

The load-ahead option instructs the sample manager to aspirate the next sample in a sample list while a current sample is running, thus eliminating the majority of the overhead time (time in addition to the chromatographic analysis time) of an injection cycle. The Load Ahead option reduces cycle time without degrading chromatographic performance.

Restrictions:

• Load Ahead Mode is designed for use with the Partial Loop with Needle Overfill injection mode.

• The first injection of a sample set and injection sets with different methods do not utilize load ahead mode.

The following figures show a standard ACQUITY UPLC system cycle time and load-ahead cycle time.

Standard ACQUITY UPLC system cycle time definition

SM wash time

Start End

Chromatographic run time

SM sample preparationand positioning

Sample injection

Systemsetup

Standard injection mode and first injection in load ahead mode

Total ACQUITY UPLC system cycle time

40

ACQUITY UPLC system load-ahead cycle time definition

Tip: The buffer delay is a “wait time” that compensates for variations in the time it takes to load a sample.

Because the first injection of each injection series does not use the load ahead injection option, the time savings depends on the number of consecutive injections made.

If the wash time and the sample prep time are equal to or greater than the chromatographic run time then load ahead does not improve the cycle time.

Loop offline option

When using the full loop injection mode or the partial loop pressure assist injection mode, choose the Loop Offline option to specify an interval, in minutes, following an injection, where the needle and sample loop return to the load position.

The Loop Offline option can also reduce the contribution of the sample loop to the dwell volume, if the loop is taken offline before the first change in the gradient. The dwell volume, however, is significant only in systems that use sample loops of volumes greater than 20 µL.

The Loop Offline option is not available for injections using the partial loop with needle overfill mode.

EndStart

Sample injection

Total ACQUITY UPLC system cycle time

Load ahead mode after the first injection

Chromatographic run time

SM sample preparationSM wash time

Buffer delaySystem setup and sample positioning into loop

Using the SM-FL 41

Determining the correct time to take the loop offline

Solutes that deposit in the sample loop because of poor solubility are not transferred to the column until the gradient composition dissolves the sample and flushes it onto the column. At that point, the solute's high retention factor (k') causes it to elute from the column in a single column volume. Choosing the correct time to take the sample loop offline ensures that all of the peaks are flushed from the loop.

If you are taking the loop offline before the first gradient change, flush it with at least five loop volumes of solvent at the initial gradient composition to completely flush the sample onto the column. In this case, use the following formula to calculate the loop offline time:

Example: For a system fitted with a 20-µL loop, in which the flow rate is 100 µL/min, the loop-offline time must be at least 1 minute.

If you are taking the sample loop offline after the end of the gradient, ensure it is completely filled with solvent of the initial gradient composition before taking it offline.

In addition, observe these considerations when determining the correct loop-offline time:

• If the strong wash volume has been significantly increased, the washing process is sometimes unfinished when the loop switches back. If such is the case, extend the loop-offline time.

• The loop offline time must not occur if the contents of the sample loop are of a higher concentration than the initial gradient conditions.

Loop offline time (min) =Nominal loop volume (μL)

Flow rate (μL/min)× 5

Loop offline time (min) =20 (μL)

100 (μL/min)× 5

Loop offline time (min) = .2 min × 5

Loop offline time (min) = 1 min

42

To choose the load-ahead and loop-offline options:

1. In the instrument method editor, click the ACQ-FL tab and then the General tab.

2. Select the boxes for the load-ahead and loop-offline options.

3. Specify a time interval for the loop-offline option, if necessary.

Tip: In the SM-FL, a programmed gradient typically flows through all parts of the instrument that contact the sample. If you initiate the loop-offline option before the gradient reaches its final conditions, the highly organic portion of your gradient does not pass through the sample loop. As a result, the gradient can fail to remove all sample from the sample loop – resulting in low sample recovery and an increased risk of carryover.

Reducing carryover

In a chromatographic system, any substance that creates unwanted peaks or excessive background noise is contamination. Carryover, a specific type of contamination, occurs when sample material remaining in the system after an injection appears as peaks in subsequent injections, compromising quantification. To optimize system performance, carryover must be minimized and held to an acceptable level (often, below the limits of detection).

Carryover can result from incorrectly installed tubing, fittings, or other hardware or by selecting ineffective wash solvents. Take these actions to reduce carryover:

• Restrict sample loop usage to one system. When reinstalling the loop, adopt the same orientation as the original installation. A reversed sample loop can cause carryover when reinstalled.

• Ensure all high and low pressure tubing connections are properly set. Tubing must seat properly (that is, without gaps) in its fittings before you tighten them. Poorly seated connections create unnecessary space-reservoirs that retain sample, increasing carryover.

• When possible, partially fill sample wells or vials to minimize sample contact with the piercing needle.

• Ensure the needle is free of debris or residue.

• Inspect and clean the tubing that connects the piercing needle to the wash pump. During wash procedures, the tubing carries drawn wash solvent through the piercing needle. If it becomes clogged, the piercing needle goes unwashed.

Using the SM-FL 43

• Ensure the sample needle and air sensor tube are clean. Over time, they can become coated with sample to the extent that wash solvents fail to clean them adequately.

• Inspect the injection port and its O-ring for sample residue, which can cause carryover. When necessary, replace those components.

• Replace the injection valve at regular, preventive-maintenance intervals. Over time, the fittings and seals can become worn, causing carryover.

• Characterize the loop and needle volume after you change the method or solvents. A useful rule of thumb is to characterize the loop and needle volume at the start of each day.

• Avoid plate or vial sealing systems that use sticky substances, which can cause carryover.

• Follow the guidelines on page 20 when selecting wash solvents.

• Add the Cycle Inject Valve event to the method. The event actuates the injection valve during a run, pushing any residual material trapped within it into the sample loop and, at the end of the gradient, injecting that material.

Diagnostic tests

You can select the sample syringe leak test, which verifies the sample path is free of leaks, from the SM-FL’s Maintain menu.

The Maintain menu also lists these functions:

• Calibrating the needle’s Z-axis, which calibrates the vertical position of the needle.

• Disabling motors, which you do before manually moving the sample tray and R-carriage.

• Parking the sample needle and injection valve, which you do before storing the system, or replacing a needle or valve.

• Replacing the needle, sample syringe, and wash syringes.

See also:

• Binary Solvent Manager Operator’s Overview and Maintenance Information document for information on the solvent manager’s leak test.

• ACQUITY Console online Help for additional information about running diagnostic tests.

44

Resolving leak sensor errors

The SM-FL can be fitted with two leak sensors, bottom and top: called the SM-FL leak sensor and the column heater leak sensor, respectively.

After approximately 1.5 mL of liquid accumulates in the leak sensor reservoir, an alarm sounds indicating that the leak sensor detected a leak.

Required materials

• Cotton swabs


• Nonabrasive, lint-free wipes

To resolve a leak sensor error:

1. In the ACQUITY UPLC Console’s Leak Sensors dialog box, determine which of the SM-FL’s 2 leak sensors detected a leak.

2. If the message reads “Leak Detected”, locate the source of the leak, and make the repairs necessary to stop it. If you need additional information, see the Column Compartments Operator’s Overview and Maintenance Information document.


Caution: To avoid scratching or damaging the leak sensor• do not allow buffered solvents to accumulate and dry on it.• do not submerge it in a cleaning bath.

Using the SM-FL 45

3. Remove the leak sensor from its reservoir, grasping it by its serrations, and pull upward on it.

Tip: If you cannot easily manipulate the leak sensor after removing it from its reservoir, detach the connector from the front of the device (see page 53).

4. Use a nonabrasive, lint-free wipe to dry the leak sensor prism.

Caution: To avoid damaging the leak sensor, do not grasp it by the ribbon cable.

Serrations

TP02891

Prism

Lint-free wipe

46

5. Roll up a nonabrasive, lint-free wipe, and use it to absorb the liquid from the leak sensor reservoir and its surrounding area.

6. With a cotton swab, absorb any remaining liquid from the corners of the leak sensor reservoir and its surrounding area.

Rolled up lint-free wipe

Leak sensor reservoir

Cotton swab

Leak sensor reservoir

Using the SM-FL 47


8. If you detached the connector from the front of the instrument, reattach it.



TP02892

T-bar



48

Maintaining the SM-FL

Contacting Waters technical service

If you are located in the USA or Canada, report malfunctions or other problems to Waters Technical Service (800 252-4752). Otherwise, phone the Waters corporate headquarters in Milford, Massachusetts (USA), or contact your local Waters subsidiary. The Waters Web site includes phone numbers and e-mail addresses for Waters locations worldwide. Visit www.waters.com.

When you contact Waters, be prepared to provide this information:

• Error message (if any)

• Nature of the malfunction

• Instrument serial numbers (see page 49)

• Flow rate

• Operating pressure

• Solvent(s)

• Detector settings (sensitivity and wavelength)

• Type and serial number of column(s)

• Sample type

• Data application version and serial number

• ACQUITY UPLC system workstation model and operating system version

For complete information on reporting shipping damages and submitting claims, see the document Waters Licenses, Warranties, and Support Services.

Locating system serial numbers

Each system instrument or device bears a serial number that facilitates service and support. Serial numbers also provide a way to create single log entries for each instrument so that you can review the usage history of a particular unit.

Be prepared to provide the serial numbers of the instruments in your system when you contact Waters customer support.

Maintaining the SM-FL 49

To view instrument information:

1. In the ACQUITY UPLC Console, select an instrument from the system tree.

2. Click Configure > View module information.

The Module Information dialog box displays this information:

• Serial number

• Firmware version

• Firmware checksum

• Component software version

Alternatives:

• From the main window, hover the pointer over the visual representation of the system instrument or device you want information about.

• Obtain the serial number from the printed labels on the device’s rear panel or inside the sample compartment door.

Maintenance schedule

Waters recommends that you perform the following routine maintenance on the SM-FL to ensure reliable operation and accurate results. When using the system throughout the day (and on nights and weekends), or when using aggressive solvents such as buffers, perform these maintenance tasks more frequently.

Recommended routine maintenance schedule:

Maintenance procedure Frequency For information...

Replace the leak sensor As needed See page 52.

Replace the sample needle When its capacity or material compatibility is inappropriate for your chromatographic need

See page 55.

50

Maintenance considerations

Safety and handling

Observe these warning and caution advisories when you perform maintenance operations on your system.

Replace the puncture needle cartridge

When you replace the sample needle or During scheduled routine maintenance

See page 55.

Calibrate the needle’s z axis After replacing the sample needle

See page 63.

Replace the sample loop When its capacity is inappropriate for your chromatographic need

See page 64.

Replace the sample syringe During scheduled routine maintenance or as needed

See page 67.

Replace the wash syringes During scheduled routine maintenance or as needed

See page 70.

Replace the injection valve cartridge

During scheduled routine maintenance or as needed

See page 74.

Clean the instrument with a soft, lint-free cloth, or paper dampened with water

As needed See page 76.

Warning: To avoid the harmful effects of personal contact with solvents, including inhalation, observe Good Laboratory Practice when you handle them. See the Material Safety Data Sheets for the solvents you use.

Recommended routine maintenance schedule: (Continued)

Maintenance procedure Frequency For information...


Proper operating procedures

To ensure your system runs efficiently, follow the procedures on page 9.

Configuring maintenance warnings

Maintenance counters provide real-time usage status information that can help you determine when to schedule routine maintenance for specific components. You can set usage thresholds and maintenance warnings that alert you when a component reaches the designated threshold limit. By setting threshold limits and monitoring these usage counters regularly, you can minimize unexpected failures and unscheduled downtime during important work. For information on setting maintenance warnings, consult the ACQUITY UPLC Console online Help.

Replacing the leak sensor

Required materials


• New leak sensor

Warning: To avoid electric shock, do not remove the instrument’s protective panels, except when replacing the wash syringes.



52

To replace the leak sensor:


2. Press down on the tab to detach the leak sensor connector from the front of the instrument.

3. Remove the leak sensor from its reservoir, grasping it by its serrations, and pull upward on it.

4. Carefully unpack the new leak sensor.

Leak sensor connector

Press down on tab to release connector

Serrations



6. Plug the leak sensor connector into the front of the SM-FL.



TP02892

T-bar



54

Replacing the sample needle

Required materials

• T6 TORX driver

• T10 TORX driver

• T20 TORX driver

• 1/4-inch open-end wrench


• Needle assembly kit

To replace the sample needle:

1. In the ACQUITY UPLC Console, select Sample Manager-FL from the system tree.

2. Remove the sample plate, if a plate is loaded.

3. Click Maintain > Replace > Needle.

Result: Doing so invokes a wizard that moves the sample needle assembly to an accessible position.

4. Open the sample compartment and fluidics compartment doors.



5. Using the T20 TORX driver, loosen the two captive screws that secure the access panel.

6. Carefully grasp the needle tubing, and hold it securely while you remove the panel.

7. Unscrew the needle assembly’s fitting from injector port 3.

Captive screw (2)Access panel

Needle assembly fitting (port 3)

Grasp tubing here when removing access panel

56

8. Hold the needle tubing, and use the TORX T10 driver to loosen the screw on the needle-tubing clamp.

9. Remove the needle tubing from the slot in the clamp assembly.

10. Hold the fitting end of the needle tubing, and push the needle latch back, to release the needle mounting cylinder from its mounting cavity.

Needle-tubing clamp

Needle latch

Mounting cavity


11. Lift the needle tip from the puncture needle cartridge at the bottom of the needle mechanism.

12. Using the T6 TORX driver, loosen the puncture needle cartridge set screw, and then remove the puncture needle cartridge.

13. Install the new puncture needle cartridge, and tighten the set screw.

Warning: To avoid puncture wounds, do not press or otherwise touch the needle tip.

Caution: To avoid damaging the needle, do not press or otherwise touch its tip.

Caution: To avoid damaging the needle tubing, and to ensure the needle fits correctly in the device, do not bend or in any way deform the needle.

Puncture needle cartridge

Needle

Puncture needle cartridge set screw

58

14. Holding the new needle assembly with both hands to control its position within the sample compartment, insert the needle’s tip into the puncture needle cartridge at the bottom of the needle mechanism.

15. Insert the needle mounting cylinder into the mounting cavity on the carriage assembly.

16. Push the needle latch downward, to secure the needle assembly.

Insert needle tip here

Needle latch


17. Route the needle tubing into the slot on the backside of the latch.

18. Carefully bend the needle tubing until it snaps into place underneath the retainer on the needle latch.

19. Bring the needle tubing up toward the slot in the clamp assembly.

Tip: The tubing bends, to accommodate the routing.

Tubing slot

Needle tubing retainer

Needle latch

60

20. Ensure that the tubing is routed as pictured below.

21. Route the needle tubing through the slot in the clamp assembly.

Caution: To prevent damage to the needle tubing, do not force it into position.


22. Ensure that the edge of the black tubing sleeve aligns with the left edge of the slot, as shown below.

23. Using the T10 TORX driver, tighten the screw on the needle-tubing clamp.

24. Insert the needle tubing fully into port 3 on the injection valve, with the two-piece ferrule assembly bottomed in the port, and then thread the fitting into the port.

25. Tightening the fitting by hand, and then add 1/4-turn using the wrench.

26. Reinstall the access panel, ensuring that the needle tubing is routed through the slot on the side of the access panel.

27. Use the T20 TORX driver to tighten the two screws that secure the access panel to the front of the unit.

28. Close the sample compartment and fluidics compartment doors.

Caution: To avoid damage to the needle during operation, allow the needle to release any residual tension before you tighten the clamp.

Edge of black tubing sleeve aligned with edge of carriage slot

TORX T10 needle clamp screw

TP03430

Access panel slot

Needle tubing

62

29. Calibrate the needle’s Z axis (see page 63).

30. Characterize the needle seal (see the ACQUITY UPLC online Help).

Calibrating the needle’s z axis

You must calibrate the needle’s z axis whenever you replace the sample needle. Failing to calibrate the needle can damage it. The calibration procedure is the same for all needles.

To calibrate the needle’s z axis

1. Open the sample manager door.

2. Remove the plates from the trays.

3. In the console, select Sample Manager > Control > Reset SM.

Result: The mechanism moves to the home position, with the needle above the wash station.

4. Click Maintain > Calibrate needle Z axis.

5. Click Start, and then click OK in the confirmation window.

Tip: To easily and efficiently achieve the correct needle setting, slide a business card under the needle.

6. Select 1.0 mm for displacement per keystroke.

7. Use the +Z button (Page Down key) to drive the needle down to within 1 millimeter of the tray surface.

8. Switch the displacement increment to 0.1 millimeter, and lower the needle until it is almost touching the surface of the business card.

9. Click Save > Yes, and then click Close.

10. Remove the card.

11. Specify the sample needle and syringe volumes, if they changed.

12. Characterize the needle seal (see the ACQUITY UPLC online Help).


Replacing the sample loop

Replace the sample loop when it clogs or when its capacity is inappropriate for your chromatographic need.

Required materials


• ACQUITY UPLC sample loop

To replace the sample loop


2. Use the wrench to remove the fittings on ports 1 and 4 of the injection valve.

Caution:

• To prevent bandspreading or excessive carryover, be sure to reinstall a previously installed sample loop in exactly the same way that it was installed before, so that the same end goes into the same port.

• To avoid leaks, remove the O-rings prior to installing the sample loop. The O-rings on the new sample loop hold the seals in place during shipping.

Caution: To avoid kinks, do not pull on the volume detection device (port 2) or sample needle (port 3) tubing.

64

Sample loop fittings on the injection valve

3. Remove the sample loop and its fittings.

4. If you are going to reuse the sample loop, label each end with the port it was connected to.

Tip: If you replaced the injection valve cartridge, do not install previously used sample loops on the new cartridge.

5. Place the sample loop and its fittings in a contamination-free bag.

6. Unpack the replacement sample loop and fittings.

7. Remove the O-rings from the sample loop.

Caution: To avoid leaks, remove the O-rings prior to installing the sample loop. The O-rings on the new sample loop are used to hold the seals in place during shipping.

Port 4 sample loop fitting

Sample loop

Sample needle

Volume detection device Port 1 sample loop

fitting

Inlet from pump

Outlet to column


8. Ensure the 2-piece ferrule is in place on one end of the sample loop. If not, slide a 2-piece ferrule and compression screw onto one end of the sample loop, and then seat the end into injection valve port 1.

9. Finger-tighten the fitting and then add 3/4-turn using the wrench.

See also: page 16.

10. Remove the fitting and inspect the ferrule to ensure it has not moved.

11. Repeat step 8 through step 10 for the other end of the sample loop and for injection valve port 4.

12. Reinstall the sample loop fitting into valve ports 1 and 4.

13. Close the fluidics compartment door.

14. Ensure the sample compartment door is closed.

15. In the ACQUITY UPLC Console, select Sample Manager from the system tree.

16. Click Maintain > Calibrate System Volume to characterize the new system volumes.

Caution:

• To prevent dead volume in the system, ensure that the ends of the loop are fully inserted into the ports of the injection valve before swaging the ferrules.

• Use only Waters approved ACQUITY UPLC loops with ACQUITY UPLC fittings. Each loop must be fitted to its injector.

Caution:

• To avoid kinks, do not push on the volume detection device (port 2) or sample needle (port 3) tubing.

• To prevent bandspreading or excessive carryover, be sure to reinstall the sample loop in exactly the same way that it was installed before, so that the same end goes into the same port.

Caution: Be careful not to crush the loop or volume detection device line.

66

Replacing the sample syringe

Recommendation: Perform the sample syringe leak test whenever you replace the sample syringe (see the ACQUITY UPLC online Help).

Air bubbles in the sample syringe adversely affect system pressure, baseline, volume, and peak area.

Replace the sample syringe when any of these conditions arise:

• The syringe plunger tip becomes worn or discolored

• You want to change to the other syringe size

• The syringe leaks or causes air bubbles

• The syringe fails the leak test

Required materials

• Degassed, weak wash solvent


• Replacement sample syringe

To replace the sample syringe:


2. Click Maintain > Replace > Sample syringe > OK.


Caution: To avoid breaking the sample syringe, do not attempt to remove air bubbles from the syringe by tapping on it.

Warning: To avoid a pinch injury or puncture wound, ensure that no injection is in progress or pending before you remove the sample syringe.


Result: Doing so invokes a wizard that moves the syringe to the “down” position.


4. Remove the knurled screw that holds the sample syringe to the syringe mounting bracket.

Sample syringe assembly components:

5. Unscrew the sample syringe, counterclockwise, until it separates from the sample syringe valve.


Sample syringe

Knurled screw

Knurled collar

68

6. Depress the syringe barrel to clear the top mounting bracket, and remove the syringe.

7. Carefully unpack the replacement sample syringe.

8. Partially fill the new syringe (by hand) with weak wash solvent (to help remove air bubbles).

Requirement: Make sure all air bubbles are removed.

9. Retract the syringe plunger so that the plunger end slides over the threaded post on the syringe guide mounting bracket.

10. Screw the new sample syringe partially into the sample syringe valve.

11. Finger-tighten the sample syringe.

12. Install and finger-tighten the knurled screw that holds the sample syringe plunger to the mounting bracket.

Caution: To avoid breaking the syringe, do not grasp it by its glass barrel. Always grasp the syringe by its knurled collar.


Mounting bracket

Threaded holder

Syringe plunger

Knurled collar


13. Run the Prime Sample Syringe Only option until no air bubbles remain in the sample syringe.


Replacing the wash syringes

Replace the wash syringes when the syringe plunger’s tip leaks, becomes worn or discolored, or fails a leak test.

To ensure accurate results, use only syringes approved by Waters for use in the ACQUITY UPLC system. The only wash syringe size available is 2.5 mL.

Required materials

• Degassed wash solvents

• Replacement wash syringes

• T20 TORX driver


To replace the wash syringes:

1. Remove the wash solvent lines from the solvent bottles.

2. Remove the two TORX T20 screws that fasten the instrument’s side panel.

3. Slide the side panel back about 2.5 cm, tilt the bottom of the panel to disengage the top lip from the instrument chassis, and then remove the panel.

Warning: To avoid puncture wounds or other injury, ensure that no injection is in progress or pending before you remove the syringe.

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Warning: To avoid injury from electric shock, do not contact any of the components in the electrical hazard area shown below.

Wash syringe assembly Electrical hazard area


4. Unscrew the weak and strong wash fittings, and then move the wash lines away from the seal wash assembly.

5. Using the 1/2-inch open-end wrench, remove the nuts that secure the wash syringes to the upper mounting bracket.

6. Loosen the knurled screws that secure the wash syringe plungers to the wash syringe mounting bracket.

7. Unscrew each syringe until it separates from the mounting bracket.

8. Pull down the syringe barrel to clear the upper mounting bracket, and remove the syringe.

9. Unpack each replacement wash syringe.

Caution: To avoid breaking the syringe, do not hold it by its glass barrel. Instead, hold the syringe by its knurled collar.

Wash syringes

Strong wash line Weak wash

line

Knurled screws

Strong wash fitting

Weak wash fitting

1/2-inch nuts

Upper mounting bracket

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Wash syringe

10. Partially fill each new wash syringe (by hand) with weak or strong wash solvent (to help remove air bubbles and to lubricate the syringe plunger).

11. Slide the plunger end of the new wash syringe over the threaded post on the syringe guide mounting bracket.

12. If necessary, depress the syringe barrel to clear the upper mounting bracket, and then screw each new wash syringe into the mounting bracket.

Tip: You sometimes must pull upward on the syringe barrel as you screw the wash syringe into the mounting bracket.

13. Using the 1/2-inch open-end wrench, install the nut that secures the wash syringe to the upper mounting bracket.

14. Install and finger tighten the knurled screw that holds each wash syringe barrel to the mounting bracket.

15. If plugs are fitted in the wash syringe tips, remove them.

16. Reattach the weak-wash and strong-wash lines, connecting the fittings finger-tight.

17. Prime the system until no bubbles appear in the syringes.

18. Grip the device’s side panel by its bottom edges.

19. Position the side panel it so that it extends approximately 2.5 cm beyond the rear of the device, with the top edge of the panel beneath the device’s top cover.

Plunger end

Syringe barrel

Knurled screw


20. Tilt the bottom of the side panel inward, and slide the side panel forward until it meets the front of the device.

Tip: The holes in the panel align with the screw holes in the chassis.

21. Secure the side panel using the two TORX T20 screws.

Replacing the injection valve cartridge

Requirement: When you replace the injection valve, you must also replace the sample loop.

Required materials

• 2-mm Allen wrench


• Injection valve cartridge

To replace the injection valve cartridge:


2. In the SM-FL information window, click Maintain > Park needle and inject valve > Yes.

3. Power-off the SM-FL.




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5. Remove the fittings attached to the injection valve cartridge.

6. Use the 2-mm Allen wrench to remove the screw at the 10 o’clock position on the injection valve cartridge.

7. Remove the injection valve cartridge from the injection valve assembly.

8. Carefully unpack the replacement injection valve cartridge.

9. Ensure that the groove in the cartridge housing and the groove on the drive clamp align.

Tip: If they do not align, turn the drive clamp until the grooves line up.

10. Insert the new injection valve cartridge into the injection valve assembly.

Note: If the injection valve cartridge does not slide fully into the injection valve assembly, contact your Waters service representative.

11. Insert the 2-mm Allen screw at the 10 o’clock position on the injection valve cartridge, and tighten it.

Sample loop

To column

From solvent manager

Volume detection device

Needle

Sample loop

Drive clamp

Aligned grooves

Injection valve cartridge housing


12. Reattach all fittings, ensuring the tubes bottom in their fitting holes.


14. Power-on the SM-FL.

Cleaning the instrument’s exterior

Clean surfaces of the SM-FL using only a soft, lint-free paper or cloth dampened with water.

Observe these requirements when cleaning device surfaces:

• Always ensure the electrical power to the device is interrupted.

• Always use eye and hand protection during the cleaning process.

• Apply the water to a clean cloth only, and then wipe the device.

• Never spray or apply the water directly onto any device surface.

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