Abstract
A simple and fast procedure for obtaining labeled monoclonal antibodies (mABs) from hybridoma culture fluid in amounts sufficient for selection of antibody pairs in a sandwich ELISA and for mAB affinity assessment has been developed. The procedure is based on a novel approach involving the binding of mABs present in hybridoma culture fluid to antigen molecules adsorbed onto an 48-well plate and the subsequent temporary biotinylation of the complexes formed. The example of mABs targeting a recombinant vaccinia virus protein A27L was used to show that the amount of labeled antibodies bound to the well is sufficient for testing 25 potential partners in a sandwich ELISA setup. The technique allows for affordable and efficient selection of mAB pairs without the need for large amounts of culture or ascites fluid and without the requirement for antibody isolation.
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Abbreviations
- mAB:
-
monoclonal antibody
- rA27L:
-
recombinant vaccinia virus protein A27L
- PBS:
-
phosphate-buffered saline
- pfu:
-
plaque-forming unit
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Original Russian Text © M.M. Zaripov, G.V. Afanas’eva, K.A. Glukhova, Yu.A. Trizna, A.S. Glukhov, I.P. Beletskii, O.V. Prusakova, 2015, published in Bioorganicheskaya Khimiya, 2015, Vol. 41, No. 4, pp. 411–415.
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Zaripov, M.M., Afanas’eva, G.V., Glukhova, K.A. et al. A simple novel procedure for the production of labeled monoclonal antibodies in analytical amounts for antibody pair screening in sandwich ELISA. Russ J Bioorg Chem 41, 364–367 (2015). https://doi.org/10.1134/S1068162015030127
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DOI: https://doi.org/10.1134/S1068162015030127