Optimization and Validation of a Multiplex, Electrochemiluminescence-Based Detection Assay for the Quantitation of Immunoglobulin G Serotype-Specific Antipneumococcal Antibodies in Human Serum

Marchese, Rocio D.; Puchalski, Derek; Miller, Pamela; Antonello, Joseph; Hammond, Olivia; Green, Tina; Rubinstein, Leonard J.; Caulfield, Michael J.; Sikkema, Daniel J.

doi:10.1128/cvi.00415-08

Cited by 80 publications

(55 citation statements)

References 20 publications

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“…It is generally typical that the vendor will provide the data supporting the lack of crosstalk. It is recommended that the feasibility test of crosstalk should be evaluated regardless of data availability from the vendor (13). One way to test for cross-talk is to vary the concentration of one analyte over the full dynamic range of the assay while keeping the other analytes in the multiplex at a low constant concentration.…”

Section: Challenges Of Cross-talkmentioning

confidence: 99%

Recommendations for Use and Fit-for-Purpose Validation of Biomarker Multiplex Ligand Binding Assays in Drug Development

Jani

Allinson²,

Berisha

et al. 2015

AAPS J

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Abstract. Multiplex ligand binding assays (LBAs) are increasingly being used to support many stages of drug development. The complexity of multiplex assays creates many unique challenges in comparison to single-plexed assays leading to various adjustments for validation and potentially during sample analysis to accommodate all of the analytes being measured. This often requires a compromise in decision making with respect to choosing final assay conditions and acceptance criteria of some key assay parameters, depending on the intended use of the assay. The critical parameters that are impacted due to the added challenges associated with multiplexing include the minimum required dilution (MRD), quality control samples that span the range of all analytes being measured, quantitative ranges which can be compromised for certain targets, achieving parallelism for all analytes of interest, cross-talk across assays, freeze-thaw stability across analytes, among many others. Thus, these challenges also increase the complexity of validating the performance of the assay for its intended use. This paper describes the challenges encountered with multiplex LBAs, discusses the underlying causes, and provides solutions to help overcome these challenges. Finally, we provide recommendations on how to perform a fit-forpurpose-based validation, emphasizing issues that are unique to multiplex kit assays.

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Section: Challenges Of Cross-talkmentioning

confidence: 99%

Recommendations for Use and Fit-for-Purpose Validation of Biomarker Multiplex Ligand Binding Assays in Drug Development

Jani

Allinson²,

Berisha

et al. 2015

AAPS J

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“…The adequacy of the specific vaccine response can be assessed by measuring postvaccination levels of antipneumococcal antibodies and by comparing these levels against predetermined cut points for either the absolute antibody level or the fold change relative to the baseline value. Measurements can be performed by a variety of analytical methods, including enzyme-linked immunosorbent assays (ELISAs) for total pneumococcal antibody level (1), ELISAs for antibodies against specific pneumococcal serotypes (2), and multiplex assays which measure the levels of a panel of serotype-specific antibodies (3)(4)(5). Because the vast majority of assays used for this purpose are lab-developed tests (LDTs) that were created and characterized by individual laboratories, the potential for interlaboratory variation in results exists.…”

mentioning

confidence: 99%

Multilaboratory Assessment of Threshold versus Fold-Change Algorithms for Minimizing Analytical Variability in Multiplexed Pneumococcal IgG Measurements

Daly

Pickering

Zhang

et al. 2014

Clin. Vaccine Immunol.

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dPneumococcal vaccination is frequently used to assess a patient's humoral immune function. The comparison of pre-and postvaccination levels of antipneumococcal antibodies is widely held to be the gold standard for documenting a response. However, many of the published criteria for defining an adequate response are based on assays that are no longer widely available. We compared the clinical classification of patient response by multiplex pneumococcal assays currently performed at three large reference laboratories using a variety of published criteria for defining responses in adults. The classification of responders agreed for 79% of the patients when using a threshold-based algorithm compared to 57 to 96% of the patients when using various fold-change-based algorithms. The highest rate of discordance was seen when the most stringent criteria for response were used (4-fold increase postvaccination in 70% of serotypes). The discordant samples tended to show similar patterns of response across all three assays, with small variations in the final number of serotypes converting postvaccination. We conclude that the use of published cut points for documenting response to pneumococcal vaccination can be affected by interlaboratory differences in pneumococcal assays, particularly for algorithms that require large fold changes for a response to be documented. However, the overall patterns of response were similar in virtually all samples, regardless of the assay used. P neumococcal vaccination is often used to evaluate a patient's response to polysaccharide antigens during a workup for deficiencies in antibody production. The adequacy of the specific vaccine response can be assessed by measuring postvaccination levels of antipneumococcal antibodies and by comparing these levels against predetermined cut points for either the absolute antibody level or the fold change relative to the baseline value. Measurements can be performed by a variety of analytical methods, including enzyme-linked immunosorbent assays (ELISAs) for total pneumococcal antibody level (1), ELISAs for antibodies against specific pneumococcal serotypes (2), and multiplex assays which measure the levels of a panel of serotype-specific antibodies (3-5). Because the vast majority of assays used for this purpose are lab-developed tests (LDTs) that were created and characterized by individual laboratories, the potential for interlaboratory variation in results exists. This may lead to difficulties when trying to interpret analytical results compared to the literature-based definitions for a therapeutic response to vaccination.The assessment of a patient's response when only a single (postvaccination) sample is available is usually done by comparing serotype-specific results against a "protective" threshold (6). However, the use of paired pre-and postvaccination samples is preferred because of the additional information provided for evaluating humoral immune function, which is the primary objective of these assays. Approaches for evaluating response from pa...

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“…The results of this comparison demonstrated excellent concordance between the two assay formats. For each of the seven Prevnar serotypes, the GMCs for at least 9 of the 12 QC samples fell within Ϯ40% of the published concentrations, thus meeting the preestablished WHO criteria for concordance (7). A study that compared three different multiplexed bead-based PnPs immunoassays, including a commercial Luminex Pn assay, to the WHO ELISA, also using the WHO QC reference panel, reported that these assays did not meet the WHO-established criteria for concordance (16).…”

Section: Discussionmentioning

confidence: 99%

Comparison of a New Multiplex Binding Assay versus the Enzyme-Linked Immunosorbent Assay for Measurement of Serotype-Specific Pneumococcal Capsular Polysaccharide IgG

Ashton¹,

Zhang

Antonello

et al. 2011

Clin. Vaccine Immunol.

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The measurement of serotype-specific anti-capsular polysaccharide antibodies remains the mainstay of pneumococcal (Pn) vaccine evaluation. New methods that allow the simultaneous measurement of antibodies to several antigens in small volumes of serum, and that agree well with existing techniques, are urgently required to support the increasing number of concomitant vaccines delivered in the infant immunization schedules and the use of extended-valency Pn vaccines. We therefore compared a relatively new multiplexed platform for measuring anti-Pn antibodies with the existing WHO consensus enzyme-linked immunosorbent assay (ELISA). A panel of 50 pediatric samples (34 collected after receipt of a heptavalent pneumococcal conjugate vaccine [PCV7] and 16 without PCV7) was analyzed across two different laboratories using a new multiplex electrochemiluminescence (ECL)-based detection assay developed for the quantitation of IgG serotype-specific antipneumococcal antibodies, and the results were compared to those obtained using the WHO consensus ELISA. For the seven serotypes measured, there was good agreement between the techniques and laboratories. The most notable difference was found between the ECL assay and the ELISA: concentrations tended to be higher in the ECL assay. For serotypes 6B, 9V, 18C, and 23F, the average increases in concentration ranged from 48 to 102%. However, the agreement rates on the proportions of samples with concentrations surrounding 0.35 g/ml were >82% for all serotypes tested. Agreement between the two laboratories running the ECL assay was generally good: agreement on proportions of samples with concentrations surrounding 0.35 g/ml was in excess of 92%, and agreement on average antibody concentrations was within 31%. We conclude that the Meso Scale Discovery (MSD) platform provides a promising new technique for the simultaneous measurement of antipneumococcal antibodies.

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Optimization and Validation of a Multiplex, Electrochemiluminescence-Based Detection Assay for the Quantitation of Immunoglobulin G Serotype-Specific Antipneumococcal Antibodies in Human Serum

Cited by 80 publications

References 20 publications

Recommendations for Use and Fit-for-Purpose Validation of Biomarker Multiplex Ligand Binding Assays in Drug Development

Recommendations for Use and Fit-for-Purpose Validation of Biomarker Multiplex Ligand Binding Assays in Drug Development

Multilaboratory Assessment of Threshold versus Fold-Change Algorithms for Minimizing Analytical Variability in Multiplexed Pneumococcal IgG Measurements

Comparison of a New Multiplex Binding Assay versus the Enzyme-Linked Immunosorbent Assay for Measurement of Serotype-Specific Pneumococcal Capsular Polysaccharide IgG

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