Measurement of mouse anti-phospholipid antibodies to solid-phase microspheres by both flow cytofluorometry and Alcian blue-pretreated microtitre plates in an ELISA.
Conventional solid-phase immunoassays measuring interactions between anti-phospholipid antibodies and phospholipids are generally characterized by problems of reproducibility and high levels of non-specific binding. Here we describe two immunoassays based on the use of phospholipids in the form of solid-phase microspheres to measure the presence of anti-phospholipid antibodies in sera. Following the production of antibodies in mice against liposomes containing lipid A, we show that flow cytofluorometric analysis provides a reproducible and sensitive way to detect anti-phospholipid antibodies. We also present a sensitive, rapid and reproducible enzyme-linked immunosorbent assay (ELISA) using Alcian blue pretreated microtitre plates and solid-phase microspheres as coating antigen. This ELISA permitted the detection of antibodies to 1/1000 dilution, while untreated plates gave negative results. Such modified ELISA procedures may be applicable to other types of molecule exhibiting solid-phase binding problems e.g. synthetic peptides (J. Immunol. Methods 175 (1994) 131-135).
Laakel
M
M
Department of Microbiology and Immunology, Université, de Montréal, Quebec, Canada.
Bouchard
M
M
Lagacé
J
J
D000423:Alcian Blue; D000818:Animals; D017152:Antibodies, Antiphospholipid / Q000032:analysis*; D004396:Coloring Agents; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods* / Q000706:statistics & numerical data; D005069:Evaluation Studies as Topic; D005260:Female; D005434:Flow Cytometry / Q000379:methods*; D007114:Immunization; D008050:Lipid A / Q000008:administration & dosage / Q000276:immunology; D008081:Liposomes / Q000008:administration & dosage / Q000276:immunology; D051379:Mice; D008807:Mice, Inbred BALB C; D008863:Microspheres; D010743:Phospholipids / Q000276:immunology; D012680:Sensitivity and Specificity
D017152:Antibodies, Antiphospholipid; D004396:Coloring Agents; D008050:Lipid A; D008081:Liposomes; D010743:Phospholipids; D000423:Alcian Blue