POLYCLONAL ANTIBODIES AGAINST HUMAN PLASMINOGEN KRINGLE 5
Authors: Kapustianenko L. G.
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The aim of the research was to obtain the polyclonal antibodies against a fragment of human plasminogen kringle 5 and to study their immunochemical properties. The following approaches were used: immunization of rabbits with plasminogen kringle 5, receiving of high-titer immune serum, synthesis of kringle 5-based affinity sorbent for selection of monospecific antibodies, chromatography on synthesized K 5-Sepharose, ELISA immunoblot assay.
The obtained polyclonal antibodies reacted in ELISA with a plasminogen K 5 fragment, and to a much lesser extent with mini-plasminogen, Lys-plasminogen, K 1-3 and K 4 plasminogen fragments, Glu-plasminogen, in descending order. Based on the dissociation constant determination it was found that these antibodies had high affinity to their epitopes within K 5 fragment (3.89・10–10 M), mini-plasminogen (5.46・10–9 М) and Lys-plasminogen (2.54・10–9 М), and insignificant affinity to K1-3, K4 and Glu-plasminogen. These antibodies reacted in immunoblotting assay with an isolated K 5 fragment of human plasminogen, Lys-Pg and mini-Pg, and did not react with K 5 in Glu-Pg and with K 1-3 or K 4 fragments. Thus, the obtained polyclonal antibodies were monospecific and had a high affinity to kringle 5. These antibodies can be used for the development immuno chemical and immunosensory methods for the quantitative determination of angiostatin K 5 in biological materials.