Immunopurification of human plasma kininogens |
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Authors: | Tytti Kärkkäinen |
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Institution: | Department of Biochemistry, University of Helsinki, Unioninkatu 35, SF-00170 Helsinki 17, Finland |
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Abstract: | Human plasma kininogens were purified by immunoadsorption on Sepharose columns using two different approaches, either removing protein impurities with the respective immunospecific polymers or applying an anti-kininogen-specific immunoadsorbent column. An anti-kininogen serum developed and investigated in this laboratory in earlier studies was used. This antiserum recognizes the native conformational determinants in the kininogen heavy chain, the common denominator in plasma kininogens, and reacts with three heterogeneous molecular forms of high mol. wt kininogen (mol. wts 103,000, 92,000 and 90,000) as well as with low mol. wt kininogen. Heterogeneity of kininogens was shown by SDS gel electrophoresis and immunoelectrophoresis. With the antibody-specific polymers the yield was 80-100% compared to 75% or lower when several consecutive immunoadsorption steps were applied to remove impurities. Both methods serve the purpose of preparing immunologically pure kininogens suitable for immunization. |
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Keywords: | high mol wt low mol wt SDS-PAGE sodium dodecyl sulfate-polyacrylamide gel electrophoresis mol wt PMSF phenylmethylsulfonyl fluoride PBR hexadimethrine bromide RIA radioimmunoassay EACA ?-aminocaproic acid KIU kallikrein inhibitory unit SRI single radial immunodiffusion F XIIa coagulation factor XII activated form A-NHS anti-normal human serum NHS normal human serum |
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