An anti-MuLVp30 monoclonal coating antibody is adsorbed onto a microtiter plate. MuLV core antigen (p30) present in the sample or standard binds to the antibodies adsorbed on the plate; an anti-MuLVp30polyclonal antibody is added and binds to the antigen captured by the first antibody.
Following incubation and wash steps, a HRP-conjugated secondary antibody is added and binds to the anti-MuLVp30polyclonal. Unbound HRP-conjugated secondary antibody is removed during the wash steps, and substrate solution reactive with HRP is added to the wells.
A colored product is formed in proportion to the amount of MuLV core antigen present in the sample. The reaction is terminated by addition of acid and absorbance is measured at 450 nm. A standard curve is prepared from recombinant MuLVp30 core antigen and sample concentration is then determined.
Figure 1: MuLV Core Antigen ELISA Standard Curve.
Figure 2: Purification of Recombinant MuLV p30 Protein. Lane 1: MW STDs; Lane 2: E.Coli Whole Lysate (-IPTG); Lane 3: E.Coli Whole Lysate (+IPTG); Lane 4: Crude Lysate; Lane 5: Lysate after Ni-NTA beads; Lane 6: Bead Wash; Lane 7: Elution Fraction for Recombinant MuLV p30 Standard.