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Updated protocols for the detection of Sotatercept and Luspatercept in human serum
Author(s) -
Reichel Christian,
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.2500
Subject(s) - biotinylation , antibody , immunoprecipitation , agarose , primary and secondary antibodies , chemistry , blot , chromatography , streptavidin , microbiology and biotechnology , computer science , biochemistry , biotin , immunology , medicine , biology , gene
We recently published two protocols for the detection of Sotatercept (ACE‐011, ACVR2A‐Fc) and Luspatercept (ACE‐536, ACVR2B‐Fc) in human serum. Both methods used covalently immobilized antibodies on agarose beads for immunoprecipitation and SAR‐PAGE/Western blotting for detection. Disadvantages were the relatively high amount of antibody required per sample (10 μg) and the need of a secondary antibody for the final detection. The updated protocols overcome these limitations by antigen–antibody complex formation in solution followed by capture of the complex with anti‐antibody‐coated magnetic beads. They also omit the secondary antibody incubation step by usage of biotinylated primary antibodies, which can be directly incubated with streptavidin‐HRP. Thus, the new protocols are faster, simpler, and cheaper and offer comparable sensitivities.