Sodium dodecyl sulphate-treated proteins as ligands in ELISA.
Insoluble proteins generally do not adsorb to microtitre wells and, therefore, cannot be used as antigens in enzyme-linked immunosorbent assay (ELISA). However, denaturation and solubilization with 2% sodium dodecyl sulphate (SDS) renders these proteins suitable ligands for ELISA. In quantitative ELISA using polyclonal antibodies as primary antibody, comparable results were obtained with native and SDS-denatured protein ligands. The binding of the antibodies to the SDS-treated ligands was completely inhibited by premixing the primary antibody with the corresponding native antigen. Nonspecific binding of primary and secondary antibodies to SDS-treated ligands was not observed. SDS-treated proteins are able to attach to ELISA microwells, retain their antigenic epitopes and do not engender an elevated background. The concentration of SDS-treated proteins required for coating is the same as that of the native proteins.
Lechtzier
Valeria
V
Experimental Pathology, The Hebrew University-Hadassah Medical School, POB 12272, Jerusalem 91120, Israel.
Hutoran
Marina
M
Levy
Talia
T
Kotler
Moshe
M
Brenner
Talma
T
Steinitz
Michael
M
D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D006801:Humans; D007074:Immunoglobulin G / Q000032:analysis / Q000187:drug effects; D008024:Ligands; D011506:Proteins / Q000032:analysis* / Q000187:drug effects / Q000276:immunology; D011950:Receptors, Cholinergic / Q000187:drug effects / Q000276:immunology; D012710:Serum Albumin, Bovine / Q000187:drug effects / Q000276:immunology; D012967:Sodium Dodecyl Sulfate / Q000494:pharmacology; D012995:Solubility
D007074:Immunoglobulin G; D008024:Ligands; D011506:Proteins; D011950:Receptors, Cholinergic; D012710:Serum Albumin, Bovine; D012967:Sodium Dodecyl Sulfate