Fixed protein A-containing staphylococci as solid-phase immunoadsorbents.
Staphylococcus aureus Cowan I containing protein A were rapidly fixed in hot 5% (w/v) trichloroacetic acid (TCA) for 6 min in order to remove the negatively charged cell wall polymer teichoic acid. This resulted in a stable IgG adsorbent able to bind 1.4 mg of human IgG/ml 10% (v/v) suspension for at least 5 months. This IgG binding ability is 25% less than for formalin-fixed bacteria, which were stable for at least 1 year. Preincubation of the bacterial adsorbents in 0.5% (v/v) Tween 20 after fixation prolonged stability and rendered prewashes prior to use unnecessary. IgG could be quantitatively eluted from TCA-fixed bacteria at pH 3.0, but not from formalin-fixed bacteria unless 80 mM MgCl2 was included in the acid buffer. This is explained by ionic interaction between IgG and teichoic acid at the bacterial surface at low pH. Non-specific binding of different proteins to the bacterial adsorbents was also studied and buffers for reducing this effect are suggested.
D000818:Animals; D000936:Antigen-Antibody Complex / Q000378:metabolism*; D001666:Binding Sites, Antibody; D002417:Cattle; D004306:Dose-Response Relationship, Immunologic; D005293:Ferritins / Q000378:metabolism; D006736:Horses; D006801:Humans; D007074:Immunoglobulin G / Q000378:metabolism; D007164:Immunosorbents / Q000378:metabolism*; D013205:Staphylococcal Protein A / Q000378:metabolism*; D013211:Staphylococcus aureus / Q000378:metabolism*; D013682:Teichoic Acids / Q000378:metabolism; D014238:Trichloroacetic Acid / Q000494:pharmacology
D000936:Antigen-Antibody Complex; D007074:Immunoglobulin G; D007164:Immunosorbents; D013205:Staphylococcal Protein A; D013682:Teichoic Acids; D014238:Trichloroacetic Acid; D005293:Ferritins