RESUMO
An Enzyme Linked Immunosorbent Assay (ELISA) is described for use in the determination of beta-endorphin antibody titers, as well as for the quantitation of naturally occurring levels of beta-endorphin in plasma and other bodily fluids. The ability of the assay to accommodate unpurified samples containing small concentrations of beta-endorphin was improved through the use of affinity purified antibodies in conjunction with a competitive inhibition ELISA. The problem of non-specific binding of beta-endorphin during competitive inhibition assays was circumvented through a two-step process in which the plate was first coated with BSA, followed by a second plate coating with poly-lysine (MW4000). The second coating with poly-lysine was found necessary in order to eliminate intermolecular void spaces following initial plate treatment with BSA. Following these procedures enabled quantitation of beta-EP at a level as low as 10 pmoles per microtitre plate well.
