A possible urease-based homologous ELISA for buffalo prolactin

Abstract

The feasibility of a direct binding as well as competitive ELISA using urease labelled probe is described for buffalo pituitary prolactin. Rabbit anti-buPRL serum and goat anti-rabbit IgG conjugated to urease were used as primary and secondary antibodies respectively. Assay was performed in polystyrene wells pretreated with 3-amino-propyltriethoxysilane (APTES) compound. APTES pretreatment provided a more receptive surface for prolactin (PRL). In direct binding mode the assay could detect 24 pg of PRL per well. In competition mode the detection limit was less than 0.1 ng of competitor PRL per well. Urease-based ELISA was found highly sensitive when the substrate solution containing pH indicator, bromocresol purple, was used at lower pH (4-4.2). No false positive reactions occurred in the control wells. A good parallelism between buffalo serum sample inhibition curve and the standard PRL inhibition curve was observed.