Studies on the sidedness of the human red blood cell membrane: preparation of stable azo-erythrocytes.
Para-diazobenzenesulfonic acid (DBSA), a non-penetrating compound, which reacts with proteins of the external surface of intact cells, was used for the preparation of AZO-erythrocytes. When coupling of p-diazobenzenesulfonic acid to human red blood cells was carried out in phosphate-buffered saline (PBS), the cells lysed within one hour, before the completion of the reaction. Elimination of sodium chloride from the reaction mixture allowed the completion of coupling by prolonging the survival of red blood cells in he diazonium salt from one hour to three hours. The survival of human erythrocytes could be further prolonged to seven hours by carrying out coupling in the presence of cyclic 3',5'-guanosine monophosphate (cGMP). Azo-erythrocytes prepared in the absence of cyclic GMP and washed free of the diazonium salt lysed in phosphate-buffered saline, but remained stable for hours in isosmotic phosphate buffers devoid of sodium chloride ions. Under identical conditions, azo-erythrocytes prepared in the presence of cyclic GMP remained stable for two days and were suited for studies on the functional and structural aspects of red blood cell membrane.
D000110:Acetylcholinesterase / Q000097:blood; D002463:Cell Membrane Permeability; D006152:Cyclic GMP; D003979:Diazonium Compounds; D004910:Erythrocyte Membrane / Q000032:analysis* / Q000201:enzymology; D004912:Erythrocytes / Q000032:analysis*; D005954:Glucosephosphate Dehydrogenase / Q000097:blood; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000097:blood; D006461:Hemolysis; D006801:Humans; D007202:Indicators and Reagents; D013425:Sulfanilic Acids / Q000031:analogs & derivatives
D003979:Diazonium Compounds; D007202:Indicators and Reagents; D013425:Sulfanilic Acids; D005954:Glucosephosphate Dehydrogenase; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases; D000110:Acetylcholinesterase; D006152:Cyclic GMP; C000469:diazobenzenesulfonic acid