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STLCC_CPLS Morrison 9/4/2015 Page 1
Genetic Analyzer: ABI
Maintenance,
Calibration, and Supplies
Prepared by: Bob Morrison
STLCC-CPLS, Instrumentation Specialist
Initiated: Sep 10, Last Update Sept 2015 (BAE)
ABI 3500 Genetic Analyzer
ABI @BRDG, R126B
Link to ABI 3500 User Manual…pdf (18mb)
Supplies
Jump to Spatial Cal: Maps array to camera
Jump to Spectral Cal: Dye/polymer/array changes
Jump to Performance Cal: Instrument vs. standards
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Genetic Analyzer: ABI, Service Info
STLCC_CPLS_Morrison 9/4/2015 Page 2
Dear Valued Customer,
Thank you for your recent service request on your model 3500, serial number 22117-071. The reference number for your
service call is 404199796. If this is for an on-site service visit, please expect a call from your Field Engineer within 4 business
hours to schedule an appointment.
On your next call, "Ask for TAC first!".
Request TAC (Technical Assistance Center) for immediate advanced phone support from Life Technologies Certified Service
Engineers.
If the problem requires on-site support, a TAC specialist will work with your field service engineer to get you back on line.
To reach TAC, contact Instrument Services at 1-800-955-6288 Option 3 then option 1 or email
[email protected] with your Serial number, Contact information and the problem you are experiencing.
For your reference, your current Contract is valid from 10-27-2012 thru 10-26-2013. Please note that this only reflects current
coverage and does not include future contracts already purchased. If you have any Service Contract questions you may reach
Service Sales by email at [email protected] or by phone at 800-955-6288 option 3 then Option 2 If you have already
purchased a future contract please disregard this notice.
Experience the new no-cost, no-hassle Instruments & Services Portal today at www.lifetechnologies.com/ISportal or ask your
Local Field Service Engineer about this exciting new tool for your lab.
Please contact us with any questions or concerns,
Thank you for doing business with Life Technologies,
Remote Services
phone: 1-800-955-6288 option 3 then option 1
fax: 760-930-2300
email: [email protected]
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Genetic Analyzer: ABI: Maintenance, Daily/Before Each Use
STLCC_CPLS_Morrison 9/4/2015 Page 3
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Genetic Analyzer: ABI: Maintenance, Weekly
STLCC_CPLS_Morrison 9/4/2015 Page 4
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Genetic Analyzer: ABI: Maintenance, Monthly
STLCC_CPLS_Morrison 9/4/2015 Page 5
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Genetic Analyzer: ABI: Maintenance, Quarterly, Annually
STLCC_CPLS_Morrison 9/4/2015 Page 6
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Genetic Analyzer: Calibration Runs, Checklist
• From Dashboard, go to Maintenance Wizards
– Wash pump and channels
• Install Conditioning pouch, Wash 25 minutes
• Remove Conditioning pouch, seal with parafilm and return to Cold room
• Mount Polymer pouch, select “validate”
• Select Fill Array with Polymer, then Finish
• Proceed to Calibration Options (Spatial, Spectral) on next slides
• After Calibration, If ABI is not to be used immediately remove the polymer
pouch, store in Cold room, and install a Conditioning pouch.
STLCC_CPLS_Morrison 9/4/2015 Page 7
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Genetic Analyzer: Spatial Calibration
STLCC_CPLS_Morrison 9/4/2015 Page 8
Do not open door
Spatial Calibration determines the position of the image from each capillary on the CCD array. (pg 99)
Spatial required
when:
1.Move or replace
cap array.
2. Open detector
door or move cell
3. Move or shake
instrument
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Genetic Analyzer: Evaluate Spatial Calibration Results
STLCC_CPLS_Morrison 9/4/2015 Page 9
May Reject for
another try, Seeking
even peaks around
6K
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Genetic Analyzer: Spectral Calibration, When required?
STLCC_CPLS_Morrison 9/4/2015 Page 10
Spectral calibration generates matrix for each capillary to compensate for dye overlap. Also used to
convert the 20 color data into 4,5,6 data.
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Genetic Analyzer: Spectral Cal Notes
STLCC_CPLS_Morrison 9/4/2015 Page 11
• 09/04/2015, BAE – spectral calibration failed due to >1 bad capillary (#s 1/A & 5/E), but all
other caps passed criteria (#s 2-4, 6-8) – report saved in “D:\RUNS\MAINT
DOCUMENTATION\Spectral Report 20150730 good but failed due to more than 1 bad cap.pdf”
(note 7/30/15 datestamp – system date changed to allow expired buffers)
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Genetic Analyzer: ABI, BigDye Spectral Cal Kit, PN 4404314
STLCC_CPLS_Morrison 9/4/2015 Page 12
1. Set 1.5ml block heater to 95C
2. Retrieve DS-33 Matrix Stand Std from Cold room, spin 15 sec at 3-6K RPMs in microcentrifuge
3. Resuspend 3uL of the Sequencing Install Standard (PN 4401459) with 297 uL of H-Di
Formamide (PN 4440753 or 4311320 from Cold Room)
4. Vortex thoroughly (max setting 15 sec), then Microcentrifuge 15 sec at 3-6K RPMs
5. Heat Standard at 95C for 5 minutes to denature, then chill on ice
6. Dispense 10 uL of denatured Standard mixture into 96-well ABI plate, wells A1-H1
7. Centrifuge plate with balance plate 1 minute at 3000 RPMs
8. Assemble plate and place on autosampler in ABI
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Genetic Analyzer: Spectral Cal; Prepare Standard
STLCC_CPLS_Morrison 9/4/2015 Page 13
We are using “Z”
June 2011 per BB
Note: In Spectral Cal run, You do NOT have to define plate or link the plate to an Assay as in normal
runs. The plate definition is known and the layout of Standards is As shown here.
Capillary Number/Configuration: 8 7 6 5 4 3 2 1
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Genetic Analyzer: Spectral Cal (only) Setup
STLCC_CPLS_Morrison 9/4/2015 Page 14
2. Select # Wells
3. Select Chemistry
4. Allow borrowing
5. Start-Run
Reads polymer RFID
3. Chemistry
Standard
-Select Sequencing
5. START RUN
4. Check Allow Borrowing,
one cap results and be
used for another
2. Select 96
Select A
Select “Z” dye set
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Genetic Analyzer: Performance Check with Spectral Cal
STLCC_CPLS_Morrison 9/4/2015 Page 15
4. Check this box to Keep/save
Sequenced and Calibration Data
2. Select General Sequencing
1. Under Performance, Select Seq Install Standard
3. Select 96, and A position
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Genetic Analyzer: ABI, History View at end of Cal Run
Observe Pass/Fail status of all rows
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Genetic Analyzer: Spectral Cal; Evaluation Quality Value, Cond
Number
STLCC_CPLS_Morrison 9/4/2015 Page 17
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Genetic Analyzer: Spectral Cal; Acceptance Criteria
STLCC_CPLS_Morrison 9/4/2015 Page 18
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Genetic Analyzer: Spectral Cal; Evaluate Results
STLCC_CPLS_Morrison 9/4/2015 Page 19
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Genetic Analyzer: ABI, Performance/Cal Sequence Results
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Genetic Analyzer: ABI, Maintenance, Shutdown Procedures
STLCC_CPLS_Morrison 9/4/2015 Page 21
Remove polymer and put
on Conditioning Pouch
for shutdown
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Genetic Analyzer: ABI; Reagents, Supplies, PNs
STLCC_CPLS_Morrison 9/4/2015 Page 22
BigDye® Terminator v3.1 Cycle Sequencing Kit
Part Number 4337455 Quantity/Package 100 rxn
Kit Contents :
800uL Tube of BigDye® Terminator v3.1 Ready Reaction Mix
Tube M13 (-21) Primer
Tube pGEM Control DNA
Two 1mL Tubes of 5x Sequencing Buffer. (PN 4336697)
BigDye XTerminator® Purification Kit - 2 mL (~100 - 20 µL
reactions) Part Number 4376486
BigDye XTerminator® Purification Kit is a DNA sequencing
reaction purification method that removes unincorporated
BigDye® terminators and salts that, if not removed, interfere with
sample injection and basecalling
Contents:
SAM solution: 9ml, PN 4376495
X Terminator Solution: 2ml, PN 4376492
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Genetic Analyzer: ABI, Cap Array, Polymers, Polymer Cap
STLCC_CPLS_Morrison 9/4/2015 Page 23
Analyzer Capillary Array, 36 cm, PN 4404683 (Forensics)
Analyzer Capillary Array, 50 cm, PN 4404685 (Sequence and Fragment)
POP-7 (384 samples) Short to Long Read
Sequencing and Fragments
Performance Optimized Polymer
PN: 4393708
Pouch Cap for 3500
PN: 4412619
Cap Array
PN 4404685
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Genetic Analyzer: ABI, Retainer, Base Set, Septas
STLCC_CPLS_Morrison 9/4/2015 Page 24
96-Well Septa for 3500/3500xL Genetic Analyzers PN 4412614 20 each
Septa Cathode Buffer Container, 3500 series PN 4410715 10 each
Standard Base PN 4410228 4 sets
96 ABI Prism Well Optical Reaction Plate with Barcode (code 128) PN 4306737 20 per pkg
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Genetic Analyzer: ABI, Sequence Analysis Reagents
(pg 258 ABI 3500 user manual)
STLCC_CPLS_Morrison 9/4/2015 Page 25
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STLCC_CPLS_Morrison 9/4/2015 Page 26
Genetic Analyzer: ABI, Sequencing Standards
(pg 258 ABI 3500 user manual)
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Genetic Analyzer: ABI, Fragment and HID Reagents
(pg 259 ABI 3500 user manual)
STLCC_CPLS_Morrison 9/4/2015 Page 27
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Genetic Analyze: ABI, DS-33 Matrix Standard Kit
STLCC_CPLS_Morrison 9/4/2015 Page 28
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Genetic Analazer: ABI, Pump Cleaning Kit, PN4414007
STLCC_CPLS_Morrison 9/4/2015 Page 29Hotlink to ABI Pump Cleaner Kit Protocol…. Pdf
If/when the Water Wash Wizard is ineffective in
cleaning the pump assembly, use this kit to clean the
entire system via several ports.
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Genetic Analyzer: ABI 3500 Consumables