Verifying New Reagent Lot Performance
Verifying New Reagent Lot PerformanceJulianne AddisonApplication Consultant, Siemens RSC Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.Answers for life.ObjectivesDescribe issues relating to changing reagent lotsDiscuss the reasons QC material cannot be relied on forevaluating new reagent lotsDescribe the new CLSI guideline EP26Outline steps to verify performance of a new reagent lot Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.Lot-to-Lot Variation Is a FactIn spite of quality efforts bymanufacturers, new reagent lotscan occasionally exhibit differentperformanceRegulatory and accreditationorganizations mandate verificationof new lot performanceTo date, there has not been astandard protocol to verify newreagent lot performance1 Martindale,et al., Validating New Reagents: Roadmaps through the Wilderness, LabMed, 37, 2006, 347-351 Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.3Handout 1
“We QC each new lot, isn’t that enough?”QC material does not interact with reagents exactly the same way asfresh patient samples Often QC material shows change in performance with a new reagent lotwhen no change is noted with patient samples Sometimes QC material shows no change, but patient samples doWhy? – altered matrix of the QC material Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.What’s a “matrix” ?Matrix:everything in the sample except what we are measuring§ QC sample matrix different from patient samples Matrix proteins in particular Matrix modified in manufacture§ Proficiency Testing (PT), EQA, Linearityand other manufactured samples alsohave an altered matrixDenatured Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.Matrix ModifiedPool serumRemove selected analytesCa 2Add analytes back at target concentrationsStabilize for storageGlucosePorcineLDH 2CaGlucoseLDHTSHRecombinantTSH Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.6Handout 2
Matrix effects on methodsMatrix effects maychange in newreagent lotMethods sensitiveto sample matrixExamples:Enzyme activity assaysReagents containmultiple biologiccomponentsMeasured valueimpacted by alteredsample matrixOne or morecomponents may bedifferent in a newreagent lotImmunoassaysImpact on QCresults varies by Method Lot QC material Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.Matrix InteractionsVary Between QC Materials130120QC brand A – Lot 123Result110QC brand A – Lot 125100QC brand AIA controlQC brand C9080New Reagent eviations: IA, immunoassay; QC, quality control. Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.8Managing Lot to Lot – verifying result qualityDemonstrate lack of impact using patient samplesCAP requires validation before using new lot:CHM.12900: Are new reagent lots and/or shipments validated before orconcurrent with use for patient testing ?NOTE: Good clinical laboratory science includes patient based comparisonswhen possibleTest patient samples with old and new reagent lots Does not require large numbers of samples Can evaluate based on clinical judgment Decide if lot to lot difference is significantWhat is an acceptable lot to lot difference for patient results ? Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.Handout 3
What is significant lot to lot difference?A single criteria like “Less that 10%” will not workTypically, comparison done by testing patient samples once with each lot Must take into account method imprecision Since imprecision varies by method, no single criteria will work for allShould be based on clinical significanceActualLot to LotDifferenceLot 1Lot 2Max Expected Differencebetween two results Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.Verifying Lot to Lot result qualityThe challenges Starting a new reagent lot isfrequent event Can happen at any time on any day Often there is very little of the“current” lot leftProtocol used needs to Be simple to perform when needed Require few resources or samples Provide a quick screen to detectclinically significant differences Must be done quickly Little time to find necessary patientsamples Limited resources available Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.New CLSI Guideline*EP26-A User Evaluation of Between-Reagent Lot Variation§ Intended to provide laboratories with a practical tool to screennew reagent lots for difference in performance§ Is useful within typical constraints of the clinical lab§ Designed to use minimum number of fresh patient samples Required number of samples determined by Size of the critical lot to lot difference Imprecision of the measurement procedure Goal for probability of detecting the critical difference May use as few as three patient samples Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.* 2013 CLSIHandout 4
EP26 – two part protocolSet up – needs time and thought Decisions and calculations made in advance Determines the number of samples to be tested Generally, done one time for each analyte Create summary checklist or spreadsheet for routine useActual evaluation of a new lot – simple & quick Readily performed when needed Simple protocol, simple arithmetic Clear criterion for acceptance Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.EP26 – Part 1 - Set upNeed to determine Critical Difference (CD)§ Maximum acceptable differencebetween lots§ Based on clinical use of analyte* Total Allowable Error cab be basis§ CD will be unique for each analyte§ Smaller CD requires more samples*1999 Stockholm Conference:Kenny D, Fraser CG, Hyltoft Petersen P, Kallner A. Strategies to set global analytical quality specifications inlaboratory medicine. Consensus agreement. Scand J Clin Lab Invest. 1999;59:585 Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.Determining Critical DifferenceTotal Allowable ErrorRECD§ Random Error (RE) imprecision – e.g. 2 * SD§ Critical Difference (CD) CD/SD determines number of samples that need to be tested CD/SD 3 may need 3 samples CD/SD 1 may need 22 samples Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.Handout 5
EP26 – Part 1 - Set upNeed to determine Critical Difference (CD) Medical decision concentration(s)§ May not be practical to obtain freshpatient samples that span measuringinterval§ Focus on concentrations that aremedically important§ Typically use concentration interval neardecision point§ Typically look at 1 – 3 concentrationintervals Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.EP26 – Part 1 - Set upNeed to determine Critical Difference (CD) Medical decision concentration(s) Historical precision performance§ Necessary to account for contribution ofimprecision to verification of new lot§ Three suggested sources Manufacturers documentation Historical data from routine QC testing Studies performed for this purpose§ At each concentration need SR – Repeatability (aka within run) SWRL – Within lot total precision§ Poorer precision requires more samples Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.EP26 – Part 1: Setup (cont’d)Need to determine Critical Difference (CD) Medical decision concentration(s) Historical precision performance Desired statistical powerStatistical power determinesprobability of detecting CDExample: Power 0.9 Implies 90% probability of detectionGoal for power depends on clinicaluse of results Critical analytes may warrant 0.95 For other analytes, 0.80 may beacceptableHigher power requires more samplesFor example, with all other f
Handout 1 Siemens Healthcare Diagnostics Inc. 2013 All rights reserved. Answers for life. Verifying New Reagent Lot Performance Julianne Addison
lot 13 j4002 1 lot 14 j4032 1 lot 15 j4033 1 lot 16 j4034 1 lot 17 j4035 1 lot 18 j4087 1 lot 19 j4090 1 lot 20 j4098 1 lot 21 j4104 1 lot 22 j4109 1 lot 23 j4118 1 lot 24 j4122 1 lot 25 j4133 1 lot 26 j4156 1 lot 27 k2038 1 lot 28 k2039 1 lot 29 k2042 1 lot 30 k2043 1 lot 31 k2044 1 lot 32 k2045 1
lot 13 j4002-1 lot 14 j4032-1 lot 15 j4033-1 lot 16 j4034-1 lot 17 j4035-1 lot 18 j4087-1 lot 19 j4090-1 lot 20 j4098-1 lot 21 j4104-1 lot 22 j4109-1 lot 23 j4118-1 lot 24 j4122-1 lot 25 j4133-1 lot 26 j4156-1 lot 27 k2038-1 lot 28 k2039-1 lot 29 k2042-1 lo
2. Prepare WR by mixing 50 parts of BCA Reagent A with 1 part of BCA Reagent B (50:1, Reagent A:B). For the above example, combine 50 ml of Reagent A with 1 ml of Reagent B. Note: When Reagent B is first added to Reagent A, a turbidity is observed that quickly disappears upon mixing to yield a clear, green WR.
The Biuret test uses as a reagent: Biuret reagent. For Lowry assay are used four reagents: reagent A, reagent B, reagent C and reagent D. For last method, Bradford, is used as a reagent Coomassie Brilliant Blue G-250. The absorbance was measured at a wavelength of 750nm for Lowry, 540 nm for Biuret and 595 nm for Bradford assay.
2. Lot #4- Decary Hall Lot 3. Lot #6- Petts Upper 4. Lot #7- Campus Center Rear Lot 5. Lot #12- Decary Hall Rear Lot 6. Lot #24- Business Center Lot There are also designated Faculty/Staff only parking spaces in the following lots: 1. Lot #8- Marcil Hall Front 2. Lot #10- Marcil Hall Upper Rear Lot 3. Lot #16-
5316 nickel chloride, hexahydrate, reagent (acs) 7791-20-0 nicl2 6h20 697 nickel sulfate, hexahydrate, reagent (acs) 10101-97-0 niso4 6h2o 793 potassium acetate, reagent (acs) 127-08-2 kc2h3o2 987 potassium chlorate, reagent (acs) 3811-04-9 kclo3 653 potassium chloride, reagent (acs) 7447-40-7 kcl 654 potassium iodide, reagent (acs) 7681-11 .
HORIBA Medical Instrument Cobas c701 Reagent ABX Pentra Cholesterol CP Reagent Lot(s) 015906329 Calibrator ABX Pentra MultiCal Calibrator Lot(s) 1608201 Matrix Serum Certification Date 04/11/2017 Network Laboratory Erasmus MC HORIBA Medical Instrument Cobas Integra 400/400 plus Reagent ABX Pentra Cholesterol CP Reagent Lot(s) 015906329
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