Improved quantification of a commercial enzyme-linked immunosorbent assay kit for measuring anti-MDA5 antibody

Abstract

Objectives: To compare the quantitative performance for measuring anti-MDA5 antibody titer of two enzyme-linked immunosorbent assay (ELISA) systems: an in-house ELISA and the commercial MESACUP^TM anti-MDA5 test.

Methods: Anti-MDA5 antibody titer was measured in sera from 70 patients with dermatomyositis using an in-house ELISA and the MESACUP^TM anti-MDA5 test side-by-side. For the commercial ELISA kit, serum samples diluted 1:101 were used according to the manufacturer’s protocol, but serial dilutions of sera were also examined to identify the optimal serum dilution for quantification.

Results: The anti-MDA5 antibody titers measured by the in-house and commercial ELISAs were positively correlated with each other (r = 0.53, p = .0001), but the antibody titer measured by the commercial ELISA was less sensitive to change after medical treatment, and 37 (80%) of 46 anti-MDA5-positive sera had antibody titer exceeding the quantification range specified by the manufacturer (≥150 index). Experiments using diluted serum samples revealed that diluting the sera 1:5050 improved the quantitative performance of the MESACUP^TM anti-MDA5 test, including a better correlation with the in-house ELISA results and an increased sensitivity to change.

Conclusion: We improved the ability of the commercial ELISA kit to quantify anti-MDA5 antibody titer by altering its protocol.

Keywords:

• anti-MDA5 antibody
• biomarker
• dermatomyositis
• enzyme-linked immunosorbent assay
• interstitial lung disease

Conflict of interest

M. Kuwana holds a patent on an anti-MDA5 antibody measuring system. M. Kuwana has received grant/research support from Actelion Pharmaceuticals, Chugai, Eisai, Mitsubishi-Tanabe, Ono, and Pfizer; and has participated in speaker bureaus for AbbVie, Actelion Pharmaceuticals, Astellas, Chugai, Eisai, Janssen, Japan Blood Products Organization, Mitsubishi-Tanabe, Ono, and Pfizer. A. Murakami is an employee of Medical and Biological Laboratories. The remaining authors state that they have no conflict of interest.

Additional information

Funding

This work was supported by a research grant for intractable diseases from the Japanese Ministry of Health, Labour, and Welfare.