Human B7-H6 Recombinant Rabbit Monoclonal Antibody [PSH10-20] - BSA and Azide free (Detector)

Sandwich ELISA analysis of Human B7-H6 matched pair antibodies

Elisa assay was performed by coating wells of a 96-well plate with 100 µl per well of capture antibody (HA723190) diluted in carbonate/bicarbonate buffer, at a concentration of 5ug/ml overnight at 4℃. Wells of the plate were washed, blocked with 150 µl 0.05% tween-20 1%BSA blocking buffer, and incubated with serial diluted Recombinant Human B7-H6 protein (HA210889) starting from 8000 pg/ml to 0 pg/ml and detect antibody (HA723191, Biotin, 0.2 µg/ml) for 1 hour at 30℃ with shaking. Then the plate was washed and incubated with 100 µl per well of SA-HRP for 0.5 hour at 30℃ with shaking. Detection was performed using an Ultra TMB Substrate for 10 minutes at room temperature in the dark. The reaction was stopped with sulfuric acid and absorbances were read on a spectrophotometer at 450 nm.

Interpolated concentrations of native B7-H6 in HepG2 and MCF7 extract samples based on a 1000 µg/ml extract load.

Interpolated concentration of native B7-H6 was measured in duplicate at different sample concentrations and interpolated from the B7-H6 standard curves. The interpolated dilution factor corrected values were plotted (mean +/- SD, n=2). The mean B7-H6 concentration was determined to be and 2,081 pg/mL in HepG2 cell extract, There was no detectable signal in MCF7 cell extract.

Interpolated concentrations of spiked B7-H6 in cell culture media samples.

The concentrations of B7-H6 were measured in duplicates, interpolated from the B7-H6 standard curves and corrected for sample dilution. Undiluted samples are as follows: cell culture media 50%. The interpolated dilution factor corrected values are plotted (mean +/- SD, n=2).