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Detection of Sotatercept (ACE‐011) in human serum by SAR‐PAGE and western single blotting
Author(s) -
Reichel Christian,
Farmer Letizia,
Gmeiner Günter,
Walpurgis Katja,
Thevis Mario
Publication year - 2018
Publication title -
drug testing and analysis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.065
H-Index - 54
eISSN - 1942-7611
pISSN - 1942-7603
DOI - 10.1002/dta.2346
Subject(s) - blot , immunoprecipitation , fusion protein , antibody , gene isoform , protocol (science) , chemistry , microbiology and biotechnology , computer science , biology , biochemistry , medicine , immunology , recombinant dna , gene , alternative medicine , pathology
A method for the detection of Sotatercept (ACE‐011, ACVR2A‐Fc) in human serum is presented. The method is a modification of a recently published protocol for Luspatercept (ACE‐536, ACVR2B‐Fc), another erythropoiesis stimulating fusion protein. Out of 27 tested antibodies against either the extracellular domain of ACVR2A or the full‐length protein, only 4 antibodies bound strongly enough to Sotatercept for usage with immunoprecipitation followed by SAR‐PAGE and Western single blotting. The adapted protocol allows the detection of 0.1 ng/mL Sotatercept in just 50 μL human serum. None of the 3 commercial ACVR2‐ELISAs was able to detect Sotatercept and the 2 tested surrogate proteins, even in the μg/mL range. As for Luspatercept, only IPG‐IEF/2D‐PAGE generated discrete isoforms. Due to the long serum half‐life, the SAR‐PAGE method will be able to detect Sotatercept for several weeks and will be very useful in doping testing.