sera were kept frozen at u2212 70 u00b0 c until use serum factors interfering with the binding of mab a to denatured albumin example 7 each serum at a final concentration of 10 was mixed with 1 u03bcg ml of monoclonal antibody mab ai n elisa reagent diluent for 1 h at room temperature the serum antibody mixtures 100 u03bcl well were then added to elisa plates that had been pre coated with 4 5 u03bcg ml denatured hsa in pbs and blocked with gelatin the elisa for the detection of bound murine antibodies binding to human albumin was then performed as described above elisa for the detection of endogenous human serum antibodies binding to dhsa example 9 duplicate wells in hibinding microtitre plates costar 2592 corning inc ny usa were coated with 100 u03bcl of dhsa or control hsa diluted in pbs at 4 5 u03bcg ml the plates were incubated at room temperature for 2 h the wells where then washed with wash buffer consisting of 0 05 tween 20 in pbs sigma followed by blocking for 2 h at 25 u00b0 c with 200 u03bcl 0 5 gelatin prepared from bovine skin sigma in pbs followed by washing in wash buffer 100 u03bcl of a 10 serum solution in elisa reagent diluent was added to the wells.