hid evolution™ – combination system user - life technologies

HID EVOlution™ – Combination SystemUSER BULLETIN | DECEMBER 2010

SUBJECT: Extracting DNA and Setting Up qPCR and STR PCR Reactions

Note: For safety and biohazard guidelines, refer to the “Safety” section in the PrepFiler™ Automated Forensic DNA Extraction Kit User Guide (PN 4393917C) and HID EVOlution™ – qPCR/STR Setup System Getting Started Guide (PN 4426903). For every chemical, read the SDS and follow the handling instructions. Wear appropriate protective eyewear, clothing, and gloves.

Purpose of this user bulletin

This user bulletin provides procedures for automated DNA extraction and qPCR and STR PCR reaction setup on the HID EVOlution™ – Combination System. This user bulletin supplements the procedures provided in the PrepFiler™ Automated Forensic DNA Extraction Kit User Guide (PN 4393917C) and the HID EVOlution™ – qPCR/STR Setup System Getting Started Guide.

In this user bulletin This user bulletin covers:

■ Human identification workflow. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3

■ Extracting DNA with the HID EVOlution™ – Combination System . . . . . . . . . . . 3Prepare reagents . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5Set up the carriers and labware for extraction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5Run maintenance scripts . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8Confirm DiTi sizes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8Use the correct volume of PrepFiler™ Wash Buffer B and other reagents . . . . . . . 8Select the appropriate extraction script . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9

■ Setting up qPCR and STR PCR reactions with the HID EVOlution™ – Combination System . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 10Set up the carriers and labware for qPCR/STR PCR reaction setup . . . . . . . . . . . 11Run maintenance scripts . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 12Set up the samples, reagents, and plasticware for qPCR setup . . . . . . . . . . . . . . . 12Set up the samples, reagents, and plasticware for STR PCR setup . . . . . . . . . . . . 14Select the appropriate qPCR or STR PCR script . . . . . . . . . . . . . . . . . . . . . . . . . . . 15

What’s new in this user bulletin

New for extraction

This user bulletin has been updated to include instructions for using a new wash buffer, PrepFiler™ Wash Buffer B, with the HID EVOlution™ – Combination System, and the related modifications to the HID EVOlution™ – Combination System worktable layout and new software scripts. The additional buffer and system modifications provide a more robust automated protocol and improve overall performance.

2 HID EVOlution™ – Combination System User Bulletin

Extracting DNA and Setting Up qPCR and STR PCR Reactions

For more information about these updates, refer to the PrepFiler™ Automated Forensic DNA Extraction Kit User Guide (PN 4393917C) and the HID EVOlution™ – Extraction System and HID EVOlution™ – Combination System User Bulletin: Validation of PrepFiler™ Wash Buffer B and the Related Modifications to Worktable Layout and Scripts for DNA Extraction (PN 4457144).

New for qPCR/STR PCR setup• Updated scripts for qPCR and STR PCR setup. Scripts have been updated with:

– Alternate mixing parameters for master mix preparation– For AmpFlSTR® kits that require AmpliTaq Gold® DNA polymerase (the

COfiler®, Identifiler®, Profiler Plus®, SGM Plus®, and Yfiler® kits), the reagent block requires only one tube of AmpliTaq Gold® DNA polymerase. Refer to the HID EVOlution™ – qPCR/STR Setup System Getting Started Guide for reagent block setup.

IMPORTANT! You must combine the volumes from two full 50-µL AmpliTaq Gold® DNA Polymerase tubes into one tube. The tube containing the combined volume must be one of the original AmpliTaq Gold® DNA Polymerase tubes provided with the kit. Use of another type of tube and/or use of less than 100 µL of AmpliTaq Gold® DNA Polymerase can cause liquid detection and transfer errors.

• New scripts for use with the Quantifiler® Duo DNA Quantification Kit. Refer to the HID EVOlution™ – Combination System User Bulletin: Quantifiler® Duo DNA Quantification Kit Support and Validation – New Scripts and Converter Software (PN 4445349) for more information.

3HID EVOlution™ – Combination System User Bulletin

Extracting DNA and Setting Up qPCR and STR PCR ReactionsHuman identification workflow

Human identification workflow

Figure 1 illustrates how the HID EVOlution™ – Combination System is used in the human identification (HID) workflow. The HID EVOlution™ – Combination System allows you to automate DNA extraction, qPCR reaction setup, and STR PCR amplification reaction setup on the same Tecan Freedom EVO™ instrument. The HID EVOlution – Combination System has 4 dedicated liquid-handling channels for extraction pipetting and 4 dedicated liquid-handling channels for qPCR/STR PCR setup pipetting. For details please contact your Applied Biosystems or Tecan representative.

Figure 1 Human identification workflow. The steps in bold can be automated with the HID EVOlution™ – Combination System.

Extracting DNA with the HID EVOlution™ – Combination System

To extract DNA with the HID EVOlution™ – Combination System, follow the instructions in the PrepFiler™ Automated Forensic DNA Extraction Kit Getting Started Guide (PN 4393917C) with added and modified steps. The added and modified steps are illustrated in Figure 2 on page 4 and explained on pages 5 through 9 of this user bulletin.

IMPORTANT! Refer to version C (August 2010) or later versions of the PrepFiler™ Automated Forensic DNA Extraction Kit Getting Started Guide for the latest instructions. A PDF of version C is available for download at www.appliedbiosystems.com.

Extract DNA (automated DNA binding, washing, and elution)

Quantify DNA (automated qPCR setup)

Amplify DNA (automated normalization and STR PCR reaction setup)

Perform Capillary Electrophoresis

Analyze STRs

Lyse samples and collect the lysates

http://www.appliedbiosystems.com


Extracting DNA and Setting Up qPCR and STR PCR ReactionsExtracting DNA with the HID EVOlution™ – Combination System

Figure 2 Automated DNA extraction workflow. The steps that include additions to the instructions in the PrepFiler™ Automated Forensic DNA Extraction Kit Getting Started Guide are highlighted. “NEW” indicates steps that are changed as the result of incorporating PrepFiler™ Wash Buffer B into the workflow.

Note: The added and modified steps are necessary because the HID EVOlution™ – Combination System instrument configuration and scripts are different than the dedicated HID EVOlution™ – Extraction System described in the Getting Started Guide.

Note: Run time is dependent on the number of samples. The table shows examples of uninterrupted run times for various numbers of samples.

Set Up Automation Instrument (Chapter 3)1. Perform routine maintenance.2. Set up disposable pipette tips.3. Set up reagents.4. Set up lysate, processing, and eluate plates.

Run Automated Extraction (Chapter 4)1. Set up a sample input file.2. Set up and run the script.3. Complete the run.4. View the output file and report.

Prepare Reagents and Samples (Chapter 2)1. Prepare reagents for lysis, binding, washing, and elution steps.2. Determine sample size or input amount.3. Select a lysis protocol.4. Perform lysis.

NEW before step 1: Set up the carriers and labware for extraction.Add to step 1: Run the appropriate maintenance scripts.Add to step 2: Confirm the size of disposable pipette tips used for extraction.NEW in step 3: Use the correct volumes of reagents including PrepFiler™ Wash Buffer B.

Added, Modified, and “NEW” Steps

NEW in step 2: Select the appropriate extraction script.

NEW in step 1: Prepare PrepFiler™ Wash Buffer B.

Number of samples Extraction run time

16 1 hour 40 minutes

32 2 hours 8 minutes





Prepare reagents Prepare Wash Buffer B by adding 95% ethanol to 200 mL low-TE buffer to bring the final volume to 500 mL.

Note: PrepFiler™ Wash Buffer B has a shelf life of 6 months if the container is kept closed when it is not in use. Do not transfer Wash Buffer B from the trough back to the storage container. Use new Wash Buffer B for each run.PrepFiler™ Wash Buffer B will be available in the PrepFiler™ Automated Forensic DNA Extraction Kits in 2011.

Refer to the PrepFiler™ Automated Forensic DNA Extraction Kit User Guide, “Prepare reagents for lysis, binding, washing, and elution steps” for instructions on preparing all other reagents.

Set up the carriers and labware for extraction

If the HID EVOlution™ system was last run for qPCR/STR setup, you must set up the extraction-specific carriers and labware before performing extraction:

1. If the DNA lysate is in tubes, remove the 3-position microplate carrier from Grid 7, then place six tube racks on grids 7 through 12.

Note: If the DNA lysate is in a plate, you do not need to remove the 3-position microplate carrier.

2. Set up carriers for the extracted DNA eluate:• If you want the extracted DNA eluate placed in tubes, make sure that there

are six tube racks on grids 1 through 6.• If you want extracted DNA eluate placed in a 96-well plate, make sure that

the metal plate adaptor is on grid 13, position 1.

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24

Remove the 3-position microplate carrier



3. Remove the 3-position DiTi tray carrier from grid 35, then replace it with a flat carrier and three DiTi boxes as shown below.

4. Place the magnetic particle tube block on grid 13, position 2.

IMPORTANT! Make sure that the tubes and the block containing the tubes are positioned as shown. Incorrect positioning can result in failure to pipette magnetic particles and/or collision of the Liquid Handling (LiHa) arm with the block.

5. Confirm that the 96-well magnetic ring stand is on grid 19, position 2.

Confirm that the block is positioned as shown.



6. Set up reagent troughs:

a. Remove reagent troughs from previous runs and correctly dispose of the reagents as described in the PrepFiler™ Automated Forensic DNA Extraction Kit Getting Started Guide or the HID EVOlution™ – qPCR/STR System Getting Started Guide.

b. Place new 100-mL reagent troughs on the worktable for PrepFiler™ Wash Buffer B and other reagents as shown:

Note: The trough layout described below is different from the originally validated layout. The validation of the new trough layout is described in HID EVOlution™ – Extraction System and HID EVOlution™ – Combination System User Bulletin: Validation of PrepFiler™ Wash Buffer B and the Related Modifications to Worktable Layout and Scripts for DNA Extraction (PN 4457144).

The worktable should now match the setup shown in the PrepFiler™ Automated Forensic DNA Extraction Kit Getting Started Guide PN rev C.

1. Elution Buffer trough(grid 27, position 1)

2. Wash Buffer B trough(grid 27, position 2)

3. Prepared Wash Buffer trough (grid 27, position 3)

4. Isopropanol trough(grid 25, position 1)

5. Lysate waste trough(grid 25, position 3)



Run maintenance scripts

After placing the carriers on the worktable, but before placing the samples, reagents and plasticware (DiTis, troughs, plates, and tubes), run the appropriate maintenance scripts.

Confirm DiTi sizes When placing DiTis on the worktable, make sure that the 3-position DiTi carrier on grid 29 contains three 200-µL DiTi trays.

Use the correct volume of PrepFiler™ Wash Buffer B and other reagents

When setting up reagents, follow the instructions in version C (August, 2010) of the PrepFiler™ Automated Forensic DNA Extraction Kit User Guide. Version C contains updated volumes for PrepFiler™ Wash Buffer and PrepFiler™ Wash Buffer B.

Before starting the run, if... ...then run

It is the first run of the day Combo_DailyStartUp

It is not the first run of the day Combo_Flush

When you run Combo_DailyStartUp or Combo_Flush, you see:• Air bubbles in the lines

and/or

• Intermittent flow from a DiTi cone

Combo_Flush one or more times until:• There are no visible air bubbles

and

• Flow from the DiTi cones is constant

There are one or more DiTis on the liquid handling arm (LiHa)

Combo_Drop_DiTis



Select the appropriate extraction script

When selecting a script according to the PrepFiler™ Automated Forensic DNA Extraction Kit Getting Started Guide, select the appropriate script from the table:

IMPORTANT! The large-sample (500-µL) protocols were not tested as part of the full validation studies that were performed by Applied Biosystems. The large-sample protocol was intended only for samples that are not submerged by the 300 µL of PrepFiler™ Lysis Buffer that is used in the standard protocol. If you intend to use the large-sample protocols, perform the appropriate validation studies. DNA yields from the large volume protocol may be lower than those that are obtained using the standard protocols with comparable sample input amounts.

If the sample

lysate is in...

And you want the eluted DNA

in...

If the lysis volume is 300 μL, use the script...

If the lysis volume is 500 μL, use the script...

a 96-well Spin Plate

a 96-well plate PrepFiler_plate_plateCombo_V1_SP1 PrepFiler_plate_plate500Combo_V1_SP1

1.5-mL microcentrifuge tubes

PrepFiler_plate_tubesCombo_V1_SP1 PrepFiler_plate_tubes500Combo_V1_SP1

1.5-mL micro-centrifuge tubes

a 96-well plate PrepFiler_tubes_plateCombo_V1_SP1 PrepFiler_tubes_plate500Combo_V1_SP1

1.5-mL microcentrifuge tubes

PrepFiler_tubes_tubesCombo_V1_SP1 PrepFiler_tubes_tubes500Combo_V1_SP1


Extracting DNA and Setting Up qPCR and STR PCR ReactionsSetting up qPCR and STR PCR reactions with the HID EVOlution™ – Combination System

Setting up qPCR and STR PCR reactions with the HID EVOlution™ – Combination System

To set up qPCR or STR PCR reactions with the HID EVOlution™ – Combination System, follow the instructions in the HID EVOlution™ – qPCR/STR Setup System Getting Started Guide (PN 4426903) with added and modified steps. The added and modified steps are illustrated in Figure 3 and explained on pages 11 through 15 of this user bulletin.

Figure 3 HID Workflow. The steps that include additions or modifications to the instructions in the HID EVOlution™ – qPCR/STR Setup System Getting Started Guide are highlighted.

Note: The added and modified steps are necessary because the HID EVOlution™ – Combination System instrument configuration and scripts are different than the dedicated HID EVOlution™ – qPCR/STR Setup System described in the Getting Started Guide.

Add before step 1: Set up the carriers and labware for qPCR reaction setup.Modify step 1: Run the appropriate maintenance scripts.Modify step 2: Set up the samples, reagents, and plasticware.Modify step 3: Select the appropriate qPCR setup script.

Add before step 1: Set up the carriers and labware for STR PCR reaction setup.Modify step 1: Run the appropriate maintenance scripts.Modify step 2: Set up the samples, reagents, and plasticware.New in step 3: Select the appropriate STR PCR setup script.

Added and Modified Steps



Note: Processing times on the HID EVOlution – Combination System are the same as the processing times on the four-channel HID EVOlution™ – qPCR/STR Setup System:• An automated qPCR reaction setup run for 80 samples takes approximately 1

hour. The run time may be slightly longer if the run includes automated DNA standards preparation.

• An automated STR PCR reaction setup run for 88 samples takes approximately 90 minutes. The run time may be longer, depending on the how many samples require dilution.

Set up the carriers and labware for qPCR/STR PCR reaction setup

If the HID EVOlution™ system was last run for extraction, you must set up the qPCR/STR-specific carriers and labware before performing qPCR or STR setup:

1. If necessary, remove the tube racks from grids 7-12 and replace with the 3-position microplate carrier.

2. If your extracted DNA is in tubes, make sure there are tube racks on grids 1 through 5 (for qPCR setup) or grids 1 through 6 (for STR setup).

3. Remove the three DiTi boxes and flat carrier from grid 35, then replace it with a 3-position DiTi tray carrier as shown below.

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24

Place the 3-position microplate carrier on grid 7.



4. Remove the magnetic particle tube block.

5. Remove reagent troughs from previous runs and correctly dispose of the reagents as described in the PrepFiler™ Automated Forensic DNA Extraction Kit Getting Started Guide or the HID EVOlution™ – qPCR/STR System Getting Started Guide.

The worktable should now match Tables 1 and 2 on pages 13 and 14.

Run maintenance scripts

After placing the carriers on the worktable, but before placing the samples, reagents and plasticware (DiTis, troughs, plates, and tubes), run the appropriate maintenance scripts.

Note: To avoid introducing air into the system liquid (degassed water) tubing, place the system liquid carboy at the same height as the worktable, and replenish the system liquid as needed before each run to avoid liquid levels dropping below one-quarter carboy during the run. The time needed for complete degassing may vary depending on the climate in each laboratory and geographical location. In some situations, it may take up to three days to fully degas the system liquid. We recommend that each laboratory maintain an additional carboy of fully degassed system liquid to use for replenishment.

Set up the samples, reagents, and plasticware for qPCR setup

When setting up the worktable for a qPCR reaction setup run, position the extracted DNA samples, reagents, and plasticware as shown in the table.

IMPORTANT! Make sure that the 3-position DiTi carrier on grid 29 contains two 200-µL DiTi trays (in positions 1 and 2) and one 50-µL DiTi tray (in position 3).

Before starting the run, if... ...then run

It is the first run of the day Combo_DailyStartUp

It is not the first run of the day Combo_Flush

When you run Combo_DailyStartUp or Combo_Flush, you see:• Air bubbles in the lines

and/or

• Intermittent flow from a DiTi cone

Combo_Flush one or more times until:• There are no visible air bubbles

and

• Flow from the DiTi cones is constant

There are one or more DiTis on the liquid handling arm (LiHa)

Combo_Drop_DiTis

You refilled DiTis or moved DiTi carriers Combo_Set_200tip_Position and

Combo_Set_50tip_Position‡

‡ These scripts reset the starting position for DiTi pickup to the first DiTi in the first DiTi tray. For 200-µL DiTis, the starting position is set to grid 29, site 1, tip position 1. For 50-µL DiTis, the starting position is set to grid 29, site 3, tip position 1.



Table 1 Worktable layout for qPCR reaction setup.

Legend Plasticware

Name as Shown in HID

EVOlution™ Software

Workstation Location(Grid Position, Site Position)

1 Extracted DNA samples (if sample are in 1.5-mL tubes)

S1 through S5 Grids 1 through 5

2 Reagent block Grid 7, site 1

3 Quantifiler PCR reaction plate ReactionPlate1 Grid13, site 1

4 Extracted DNA samples (if samples are in a 96-well plate)

Samples Grid 13, site 3

5 96-well magnetic ring stand and 2-position carrier (not used in qPCR setup, but remains on the worktable).

6 Te-Shake™ (not used in qPCR setup, but remains on the worktable).

7 DiTi waste

8 100-mL trough for T10E0.1 buffer TE Grid 26, second trough

9 and 10 200-µL DiTi trays 200uL-1

200uL-2

Grid 29, sites 1 and 2

11 through 14

50-µL DiTi trays 50uL-1

50uL-2

50uL-3

50uL-4

Grid 29, site 3

Grid 35, sites 1 through 3

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 4831 32

28

6

9

10

11

12

13

7

3 5

4

1



Set up the samples, reagents, and plasticware for STR PCR setup

When setting up the worktable for an STR PCR amplification reaction setup run, position the extracted DNA samples, reagents, and plasticware as shown in Table 2.

IMPORTANT! Make sure that the 3-position DiTi carrier on grid 29 contains two 200-µL DiTi trays (in positions 1 and 2) and one 50-µL DiTi tray (in position 3).

Table 2 Worktable layout for STR PCR amplification reaction setup.

Legend PlasticwareName as Shown in

HID EVOlution™ Software

Workstation Location(Grid Position, Site

Position)

1 Extracted DNA samples (if samples are in 1.5-mL tubes) S1 through S6 Grids 1 through 6

2 Reagent block Grid 7, site 1

3 and 5 Dilution plates‡

‡ Two empty 96-well plates are required. For samples that require dilution, the system pipettes the samples to the Predilution 2 plate. If additional plate wells are required for dilution, the system pipettes the samples to the Predilution 1 plate.

Predilution1 Grid 7, site 2

Predilution2 Grid 13, site 2

4 Extracted DNA samples (if samples are in a 96-well plate) Samples Grid 13, site 1

6 STR PCR reaction plate STRplate Grid 13, site 3

7 96-well magnetic ring stand and 2-position carrier (not used in STR PCR setup, but remains on the worktable).

8 Te-Shake (not used in STR PCR setup, but remains on the worktable).

9 DiTi waste

10 100-mL trough for T10E0.1 buffer TE Grid 26, second trough

11 and 12


200uL-2

Grid 29, sites 1 and 2

13 through 16


50uL-2

50uL-3

50uL-4

Grid 29, site 3

Grid 35, sites 1 through 3

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 5031 32

210

8

11

12

13

14

15

9

4 7

6

53

1



Select the appropriate qPCR or STR PCR script

When selecting a script according to the HID EVOlution™ – qPCR/STR Setup System Getting Started Guide, select the appropriate script for the HID EVOlution – Combination System from Table 3 or Table 4:

Table 3 Freedom EVOware® software script selection for qPCR setup on the HID EVOlution™ – Combination System

Table 4 Freedom EVOware® software script selection for STR PCR setup on the HID EVOlution™ – Combination System

If you are using the kit... and the sample DNA is in... ...use the script

Quantifiler® Human a 96-well plate QuantifilerHuman_plateCombo_V1_SP1.esc

1.5-mL microcentrifuge tubes QuantifilerHuman_tubesCombo_V1_SP1.esc

Quantifiler® Human Y Male a 96-well plate QuantifilerY_plateCombo_V1_SP1.esc

1.5-mL microcentrifuge tubes QuantifilerY_tubesCombo_V1_SP1.esc

Quantifiler® Human AND Human Y Male

a 96-well plate Quantifiler_Human_Y_plateCombo.esc

1.5-mL microcentrifuge tubes Quantifiler_Human_Y_tubesCombo.esc

Quantifiler® Duo a 96-well plate QuantifilerDuo_plateCombo_V1_SP1.esc

1.5-mL microcentrifuge tubes QuantifilerDuo_tubesCombo_V1_SP1.esc

If you are using the kit... and the sample DNA is in... ...use this script

Identifiler® a 96-well plate Identifiler_plateCombo_V1_SP1.esc

1.5-mL microcentrifuge tubes Identifiler_tubesCombo_V1_SP1.esc

Yfiler® a 96-well plate Yfiler_plateCombo_V1_SP1.esc

1.5-mL microcentrifuge tubes Yfiler_tubesCombo_V1_SP1.esc

MiniFiler™ a 96-well plate MiniFiler_plateCombo_V1_SP1.esc

1.5-mL microcentrifuge tubes MiniFiler_tubesCombo_V1_SP1.esc

COfiler® a 96-well plate COfiler_plateCombo_V1_SP1.esc

1.5-mL microcentrifuge tubes COfiler_tubesCombo_V1_SP1.esc

Part Number 4427707 Rev. C 12/2010

Technical Resources and SupportFor the latest technical resources and support information for all locations, please refer to our Web site atwww.appliedbiosystems.com

Headquarters5791 Van Allen Way | Carlsbad, CA 92008 USAPhone 760.603.7200www.lifetechnologies.com

For Research, Forensic, or Paternity Use Only. Not intended for any animal or human therapeutic or diagnostic use.Information in this document is subject to change without notice.

APPLIED BIOSYSTEMS DISCLAIMS ALL WARRANTIES WITH RESPECT TO THIS DOCUMENT, EXPRESSED OR IMPLIED, INCLUDING BUT NOT LIMITED TO THOSE OF MERCHANTABILITY OR FITNESS FOR A PARTICULAR PURPOSE. TO THE FULLEST EXTENT ALLOWED BY LAW, IN NO EVENT SHALL APPLIED BIOSYSTEMS BE LIABLE, WHETHER IN CONTRACT, TORT, WARRANTY, OR UNDER ANY STATUTE OR ON ANY OTHER BASIS FOR SPECIAL, INCIDENTAL, INDIRECT, PUNITIVE, MULTIPLE OR CONSEQUENTIAL DAMAGES IN CONNECTION WITH OR ARISING FROM THIS DOCUMENT, INCLUDING BUT NOT LIMITED TO THE USE THEREOF, WHETHER OR NOT FORESEEABLE AND WHETHER OR NOT APPLIED BIOSYSTEMS IS ADVISED OF THE POSSIBILITY OF SUCH DAMAGES.

NOTICE TO PURCHASER: Limited Use Label License

The products shown in this user bulletin may be covered by one or more Limited Use Label License(s). Please refer to the respective product documentation or the Applied Biosystems website under www.appliedbiosystems.com for the comprehensive license information. By use of these products, the purchaser accepts the terms and conditions of all applicable Limited Use Label Licenses. These products are sold for research, forensic, or paternity use only, and are not intended for human or animal diagnostic or therapeutic uses unless otherwise specifically indicated in the applicable product documentation or the respective Limited Use Label License(s). Further information on purchasing licenses may be obtained by contacting the Director of Licensing, Applied Biosystems, 850 Lincoln Centre Drive, Foster City, California 94404, USA.

The trademarks mentioned herein are the property of Life Technologies Corporation or their respective owners.

Freedom EVO and Freedom EVOware are registered trademarks and HID EVOlution is a trademark of Tecan Group. Excel is a trademark of Microsoft Corporation.

© 2010 Life Technologies Corporation. All rights reserved.

Profiler Plus® a 96-well plate ProfilerPlus_plateCombo_V1_SP1.esc

1.5-mL microcentrifuge tubes ProfilerPlus_tubesCombo_V1_SP1.esc

SEfiler Plus™ a 96-well plate SEfilerPlus_plateCombo_V1_SP1.esc

1.5-mL microcentrifuge tubes SEfilerPlus_tubesCombo_V1_SP1.esc

SGM Plus® a 96-well plate SGMPlus_plateCombo_V1_SP1.esc

1.5-mL microcentrifuge tubes SGMPlus_tubesCombo_V1_SP1.esc

If you are using the kit... and the sample DNA is in... ...use this script

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hid evolution™ – combination system user - life technologies

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