fx method development

Genotyping Process Genotyping Process ImprovementsImprovements

- - Focus on Biomek FX Method Development -Focus on Biomek FX Method Development -

OverviewOverview

Genotyping Assay Prep ReviewGenotyping Assay Prep Review

Process Improvement GoalsProcess Improvement Goals

Process Improvement StrategyProcess Improvement Strategy

Biomek FX Method DevelopmentBiomek FX Method Development

Process ImpactsProcess Impacts

AcknowledgementsAcknowledgements


Nuclease-freeNuclease-freeH2OH2O

2X TaqMan2X TaqManMaster MixMaster Mix

Process Improvement GoalsProcess Improvement Goals

Why improve a reliable and high-quality process?Why improve a reliable and high-quality process?

ReduceReduce direct cost of consumables and reagents direct cost of consumables and reagents ReduceReduce indirect cost of human resources indirect cost of human resources IncreaseIncrease or maintain genotype throughput or maintain genotype throughput IncreaseIncrease or maintain quality and reliability of data or maintain quality and reliability of data

ReduceReduce our operating cost our operating cost

ReduceReduce our cost per genotype our cost per genotype

IncreaseIncrease competitiveness of Genotyping Services Lab competitiveness of Genotyping Services Lab

Process Improvement StrategyProcess Improvement Strategy> > R&D EffortsR&D Efforts

NTC Amplification ExperimentNTC Amplification Experiment Towards Towards improvingimproving quality and reliability of data quality and reliability of data

Contamination ExperimentContamination Experiment Towards Towards improvingimproving quality and reliability of data quality and reliability of data

Experimental ProtocolsExperimental Protocols Towards Towards reducingreducing cost of consumables, reagents, cost of consumables, reagents,

and human resourcesand human resources Towards Towards improvingimproving genotype throughput genotype throughput

Process Improvement StrategyProcess Improvement Strategy> Experimental Pipeline Variables> Experimental Pipeline Variables

1.1. Use Protedyne Biocube to deliver 3uL reaction mixtureUse Protedyne Biocube to deliver 3uL reaction mixtureReductionReduction of reagents of reagents

2.2. Use Biomek FX as sole liquid-handling instrument to deliver 5uL Use Biomek FX as sole liquid-handling instrument to deliver 5uL reaction mixturereaction mixture

ReductionReduction of reagents, consumables, and human resources of reagents, consumables, and human resourcesImprovingImproving genotype throughput genotype throughput

3.3. Use Biomek FX as sole liquid-handling instrument to deliver 3uL Use Biomek FX as sole liquid-handling instrument to deliver 3uL reaction mixturereaction mixture

CombinationCombination of variables 1 and 2 of variables 1 and 2

4.4. Lower annealing temperature in Viper thermocycler protocolsLower annealing temperature in Viper thermocycler protocolsImprovingImproving quality and reliability of data quality and reliability of data

Process Improvement StrategyProcess Improvement Strategy> R&D Results> R&D Results

Success of Genotyping R&D effortsSuccess of Genotyping R&D efforts Better understanding of NTCsBetter understanding of NTCs Better understanding of how contamination affects our data Better understanding of how contamination affects our data

qualityquality Lowering annealing temperature improves amplificationLowering annealing temperature improves amplification Lower reaction volume of 3 uL Lower reaction volume of 3 uL does notdoes not adversely affect adversely affect

amplification and data qualityamplification and data quality Biomek FXBiomek FX is a viable candidate for consolidating our liquid is a viable candidate for consolidating our liquid

handling needshandling needs

Biomek FX Method DevelopmentBiomek FX Method Development> > Workflow ExpansionWorkflow Expansion

Experimental Protocols were Experimental Protocols were carried out using the 4SNP/4Q carried out using the 4SNP/4Q workflowworkflow

1SNP/4+Q1SNP/4+QMultiplateMultiplate

8SNP/4Q8SNP/4Q

4SNP/4Q4SNP/4Q

2SNP/4Q2SNP/4Q

1SNP/4Q1SNP/4Q

1SNP/3Q1SNP/3Q

BiomekBiomekFXFX

• Expand Biomek FX Methods Expand Biomek FX Methods to accommodate all of our lab’s to accommodate all of our lab’s workflowsworkflows

(Biomek FX) Method Development(Biomek FX) Method Development> Guidelines> Guidelines

TimeTime – – streamline individual steps to decrease total runtimestreamline individual steps to decrease total runtime

ConsumablesConsumables – ex. span-8 can replace 96-head / FX tips – ex. span-8 can replace 96-head / FX tips

PrecisionPrecision – how consistent are your results? – how consistent are your results?

AccuracyAccuracy – are you delivering the correct amount of liquid? – are you delivering the correct amount of liquid?

ContaminationContamination – are you contaminating any reagents or products? – are you contaminating any reagents or products?

Biomek FX Method DevelopmentBiomek FX Method Development> Problems encountered / resolved> Problems encountered / resolved

Improper span-8 positioning to 384-plate wells (stabbing)Improper span-8 positioning to 384-plate wells (stabbing) Framed span-8 using deck position framing tool Framed span-8 using deck position framing tool Adjusted 384-plate labware definition settings Adjusted 384-plate labware definition settings Framed span-8 to specific labware (384-plate) on each deck Framed span-8 to specific labware (384-plate) on each deck

position position

Currently exploring ways to improve liquid deliveryCurrently exploring ways to improve liquid delivery Current system (1mL syringes) can deliver Current system (1mL syringes) can deliver 5uL5uL with c.v. of with c.v. of

+/- 1uL (+/- 1uL (20%20%) upon visual inspection) upon visual inspection

Current system (1mL syringes) can deliver Current system (1mL syringes) can deliver 3uL3uL with c.v. of with c.v. of +/- 2uL (+/- 2uL (67%67%) upon visual inspection) upon visual inspection

Currently exploring advantages and disadvantages of using Currently exploring advantages and disadvantages of using 500 uL and 250 uL syringes, which may improve pipetting 500 uL and 250 uL syringes, which may improve pipetting accuracyaccuracy

Biomek FX Method DevelopmentBiomek FX Method Development> Implementation> Implementation

Nuclease-freeNuclease-freeH2OH2O

2X TaqMan2X TaqManMaster MixMaster Mix

Process ImpactsProcess Impacts

Consumables & ReagentsConsumables & Reagents

Eliminate splash lidsEliminate splash lids Eliminate OMM platesEliminate OMM plates Eliminate Axygen (FX) tipsEliminate Axygen (FX) tips Eliminate 24-well bioblockEliminate 24-well bioblock Eliminate Biocube tipsEliminate Biocube tips Eliminate OMM plate reagent dead volumeEliminate OMM plate reagent dead volume Implement 3uL protocols to reduce reaction volume by 40%Implement 3uL protocols to reduce reaction volume by 40%

InstrumentationInstrumentation

Eliminate costly $225,000 / year Protodyne service contractEliminate costly $225,000 / year Protodyne service contract Free up lab space that Biocubes currently occupyFree up lab space that Biocubes currently occupy Allow for greater flexibility and control with regards to internal R&D / Allow for greater flexibility and control with regards to internal R&D /

process improvement / process development effortsprocess improvement / process development efforts

Process ImpactsProcess ImpactsLIMS / Software ResourcesLIMS / Software Resources Reduce database storage needs by removing one container (OMM)Reduce database storage needs by removing one container (OMM) Reduce software resource time needed to maintain and test the Reduce software resource time needed to maintain and test the

intermediate SOP for new LIMS buildsintermediate SOP for new LIMS builds Reduce scanning and tracking errors by consolidating all liquid transfer Reduce scanning and tracking errors by consolidating all liquid transfer

onto one instrumentonto one instrument

Personnel Resources Reduce personnel resources for processing by 1-2 per pipeline Reduce human plate handling & transport errors by eliminating

intermediate (OMM) container

Additional Benefits Greatly reduce stockroom storage needs Eliminate 100% of recycling and 75% of trash produced by our

lab

Process ImpactsProcess Impacts> Additional Concerns> Additional Concerns

We lose initial selling point of ‘impressive’ Biocubes with We lose initial selling point of ‘impressive’ Biocubes with potential customers and lab tourspotential customers and lab tours

Need to consider a more robust plate sealing solutionNeed to consider a more robust plate sealing solution

Biomek FX wear and tear

AcknowledgementsAcknowledgements

David MerrillDavid Merrill

Cinna MonighettiCinna Monighetti

TaqMan Genotyping GroupTaqMan Genotyping Group

fx method development

Technology

use biomek fx

ul syringes

biomek fx methods

data qualitybiomek fx

head fx tipsprecision

direct cost of consumables

liquid deliverycurrent

reliability of datareduce