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The New and the Old: Platform Cross-Validation of Immunoaffinity MASS Spectrometry versus ELISA for PromarkerD, a Predictive Test for Diabetic Kidney Disease.

ABSTRACT: PromarkerD is a proteomics derived test for predicting diabetic kidney disease that measures the concentrations of three plasma protein biomarkers, APOA4, CD5L and IBP3. Antibodies against these proteins were developed and applied to a multiplexed immunoaffinity capture mass spectrometry assay. In parallel, and facilitating current clinical laboratory workflows, a standard ELISA was also developed to measure each protein. The performance characteristics of the two technology platforms were compared using a cohort of 100 samples, with PromarkerD test scores demonstrating a high correlation (R = 0.97). These technologies illustrate the potential for large scale, high throughput clinical applications of proteomics now and into the future.

SUBMITTER: Bringans S

PROVIDER: S-EPMC7709118 | biostudies-literature | 2020 Oct

REPOSITORIES: biostudies-literature

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The New and the Old: Platform Cross-Validation of Immunoaffinity MASS Spectrometry versus ELISA for PromarkerD, a Predictive Test for Diabetic Kidney Disease.

Bringans Scott S Peters Kirsten K Casey Tammy T Ito Jason J Lipscombe Richard R

Proteomes 20201028 4

PromarkerD is a proteomics derived test for predicting diabetic kidney disease that measures the concentrations of three plasma protein biomarkers, APOA4, CD5L and IBP3. Antibodies against these proteins were developed and applied to a multiplexed immunoaffinity capture mass spectrometry assay. In parallel, and facilitating current clinical laboratory workflows, a standard ELISA was also developed to measure each protein. The performance characteristics of the two technology platforms were compa ...[more]

PMID: 33126588

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Project description:Background:PromarkerD is a novel proteomics derived blood test for predicting diabetic kidney disease (DKD). The test is based on an algorithm that combines the measurement of three plasma protein biomarkers (CD5L, APOA4, and IBP3) with three clinical variables (age, HDL-cholesterol, and eGFR). The initial format of the assay used immunodepletion of plasma samples followed by targeted mass spectrometry (MRM-LCMS). The aim of this study was to convert the existing assay into an immunoaffinity approach compatible with higher throughput and robust clinical application. Methods:A newly optimised immunoaffinity-based assay was developed in a 96 well format with MRM measurements made using a low-flow LCMS method. The stability, reproducibility and precision of the assay was evaluated. A direct comparison between the immunoaffinity method and the original immunodepletion method was conducted on a 100-person cohort. Subsequently, an inter-lab study was performed of the optimised immunoaffinity method in two independent laboratories. Results:Processing of plasma samples was greatly simplified by switching to an immunoaffinity bead capture method, coupled to a faster and more robust microflow LCMS system. Processing time was reduced from seven to two days and the chromatography reduced from 90 to 8 min. Biomarker stability by temperature and time difference treatments passed acceptance criteria. Intra/Inter-day test reproducibility and precision were within 11% CV for all biomarkers. PromarkerD test results from the new immunoaffinity method demonstrated excellent correlation (R?=?0.96) to the original immunodepletion method. The immunoaffinity assay was successfully transferred to a second laboratory (R?=?0.98) demonstrating the robustness of the methodology and ease of method transfer. Conclusions:An immunoaffinity capture targeted mass spectrometry assay was developed and optimised. It showed statistically comparable results to those obtained from the original immunodepletion method and was also able to provide comparable results when deployed to an independent laboratory. Taking a research grade assay and optimising to a clinical grade workflow provides insights into the future of multiplex biomarker measurement with an immunoaffinity mass spectrometry foundation. In the current format the PromarkerD immunoaffinity assay has the potential to make a significant impact on prediction of diabetic kidney disease with consequent benefit to patients.

Dataset Information

The New and the Old: Platform Cross-Validation of Immunoaffinity MASS Spectrometry versus ELISA for PromarkerD, a Predictive Test for Diabetic Kidney Disease.

Publications

The New and the Old: Platform Cross-Validation of Immunoaffinity MASS Spectrometry versus ELISA for PromarkerD, a Predictive Test for Diabetic Kidney Disease.

Similar Datasets

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