Affinophoresis of red blood cells: Surface antigen‐specific acceleration of electrophoresis

Abstract

A method employing the technique of affinophoresis to increase the electrophoretic mobility of specific cells according to their surface antigens was developed. Red blood cells were treated consecutively with the maximum subagglutinating dose of an anti-red blood cell serum, a biotinylated second antibody, avidin and finally with a negatively charged biotin-affinophore which was prepared by coupling biotin to polylysine (average degree of polymerization, 270 or 1150), followed by complete succinylation. The electrophoretic mobility of cells was analyzed with an automatic cell electrophoresis analyzer. The use of a homologous anti-serum increased the electrophoretic mobility of rabbit, human and rat blood cells by 2.9, 1.7 and 1.6 times, respectively. A larger affinophore containing fewer biotin moieties was more effective. In the case of a mixture of red blood cells from two species, cells from only one species could be accelerated by using homologous antiserum, e.g., affinophoresis of a mixture of human and rat red blood cells by using either homologous antiserum gave two separate peaks on the histogram, whereas a single peak would be obtained in usual electrophoresis because there is little difference in the original migration velocities of the two cell types.

Lectin-induced retardation of subcellular organelles during preparative density gradient electrophoresis: Selective purification of plasma membranes

“…Envision coating desired organelles with specific antibodies (as in [42,43] and [44]) for their retardation: the electrophoretic retardation will then depend upon the number of available binding sites. Such affinity electrophoresis can be greatly enhanced and particles can be accelerated considerably by introducing thousands of anionic charges to a second antibody in the same manner as has been proposed by Shimura et al [62]. These authors coupled polylysine to a second antibody.…”