Magnetic-bead enzyme-linked immunosorbent assay verifies adsorption of ligand and epitope accessibility.
Antigen- and antibody-coated magnetic beads, which are commercially available as activated particles and are readily coated with a variety of molecules, are excellent devices for selecting specific cell populations. We describe an enzyme-linked immunosorbent assay (ELISA)-based method that validates the adsorption of ligand to magnetic beads and verifies the surface accessibility of specific epitopes of the particular ligand. Accordingly, various ligands at a wide range of concentrations were incubated with magnetic beads and adsorption was then assessed using either a specific primary antibody and a secondary alkaline phosphatase-conjugated antibody or a specific primary alkaline phosphatase-conjugated antibody. The method is straightforward and fast and, due to the very low nonspecific background binding, it requires extremely small amounts of beads and ligand. It can be easily performed on newly prepared beads before using them for selection. Magnetic-bead ELISA also confirms the accessibility of specific epitopes on the surface of the beads, which corresponds to the primary antibody used in the assay.
Kourilov
Vitaly
V
Experimental Pathology, The Hebrew University-Hadassah Medical School, POB 12272, Jerusalem 91120, Israel.
Steinitz
Michael
M
D000327:Adsorption; D016229:Amyloid beta-Peptides / Q000032:analysis / Q000276:immunology; D000937:Antigen-Antibody Reactions; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D000939:Epitopes / Q000032:analysis* / Q000276:immunology; D006514:Hepatitis B Surface Antigens / Q000032:analysis / Q000276:immunology; D006801:Humans; D018189:Immunomagnetic Separation / Q000379:methods*; D008024:Ligands
D016229:Amyloid beta-Peptides; D000939:Epitopes; D006514:Hepatitis B Surface Antigens; D008024:Ligands