Rapid procedure for coupling protein antigens to red cells to be used in plaque assays, by prewashing in chromium chloride.
A rapid and efficient procedure is described for the coupling of proteins (protein A, ovalbumin, albumin and chicken gamma globulin) to sheep red blood cells (SRBC) to be used in antigen-specific or protein A plaque assays. This modification of the original procedure has three distinct features: prewash of the red cells with a low concentration of essentially freshly prepared CrCl3, use of a relatively high concentration of CrCl3 in the reaction mixture and a coupling time of only 4 min. Protein A plaque assays performed with such target cells have the same sensitivity as those employing red cells coupled with protein A according to the original procedure. Studies with hybridomas secreting antibody specific for a protein antigen showed that antigen-specific plaque assays employing target red cells coupled with the protein antigen according to the modified procedure have the same sensitivity as the protein A plaque assay. The modified procedure greatly facilitates cellular studies on antibody formation after immunization with protein antigens.
Savelkoul
H F
HF
Department of Cell Biology, Immunology and Genetics, Erasmus University, Rotterdam, The Netherlands.
Greeve
A A
AA
Rijkers
G T
GT
Marwitz
P A
PA
Benner
R
R
D000818:Animals; D001789:Blood Group Antigens; D001854:Bone Marrow Cells; D002712:Chlorides; D002857:Chromium; D017608:Chromium Compounds; D004905:Erythrocyte Aging; D004912:Erythrocytes / Q000276:immunology* / Q000502:physiology; D005260:Female; D006462:Hemolytic Plaque Technique; D006825:Hybridomas / Q000032:analysis; D007202:Indicators and Reagents; D051379:Mice; D008807:Mice, Inbred BALB C; D010047:Ovalbumin / Q000276:immunology; D012756:Sheep; D013154:Spleen / Q000166:cytology; D013205:Staphylococcal Protein A
D001789:Blood Group Antigens; D002712:Chlorides; D017608:Chromium Compounds; D007202:Indicators and Reagents; D013205:Staphylococcal Protein A; D002857:Chromium; D010047:Ovalbumin; C017133:chromous chloride