Biotin Conjugated Human CD80 Recombinant Rabbit Monoclonal Antibody [PSH09-63] - Detector

Sandwich ELISA analysis of human CD80 matched pair antibodies

Elisa assay was performed by coating wells of a 96-well plate with 100 µl per well of capture antibody (HA723123) diluted in carbonate/bicarbonate buffer, at a concentration of 2ug/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 CD80 protein (HA210994) starting from 4000 pg/ml to 0 pg/ml and detect antibody (HA723127B, 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 CD80 in Raji and HUVEC extract samples based on a 1000 µg/ml extract load.

Interpolated concentration of native CD80 was measured in duplicate at different sample concentrations and interpolated from the CD80 standard curves. The interpolated dilution factor corrected values were plotted (mean +/- SD, n=2). The mean CD80 concentration was determined to be 40,639 pg/mL in Raji cell extract, There was no detectable signal in HUVEC cell extract.

Interpolated concentrations of spiked CD80 in cell culture media samples.

The concentrations of CD80 were measured in duplicates, interpolated from the CD80 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).