Human CD27/TNFRSF7 Recombinant Rabbit Monoclonal Antibody [PSH08-33] - BSA and Azide free (Detector)

Sandwich ELISA analysis of Human CD27 matched pair antibodies

Elisa assay was performed by coating wells of a 96-well plate with 100 µl per well of capture antibody (HA722973) diluted in carbonate/bicarbonate buffer, at a concentration of 2 µg/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 CD27 protein (HA210880) starting from 4000 pg/ml to 0 pg/ml and detect antibody (HA722975, 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.

Sandwich ELISA analysis of Human CD27 matched pair antibodies

Elisa assay was performed by coating wells of a 96-well plate with 100 µl per well of capture antibody (HA722974) diluted in carbonate/bicarbonate buffer, at a concentration of 2 µg/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 CD27 protein (HA210880) starting from 4000 pg/ml to 0 pg/ml and detect antibody (HA722975, 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 CD27 in human cell culture supernatant samples.

Interpolated concentration of native CD27 was measured in duplicate at different sample concentrations and interpolated from the CD27 standard curves. Undiluted samples were 50% cell supernatant. The interpolated dilution factor corrected values were plotted (mean +/- SD, n=2). The mean CD27 concentration was determined to be 3084 pg/ml in Daudi and 3995 pg/ml in Raji in cell culture supernatant, undetectable in MCF7 cell culture supernatant.

Interpolated concentrations of native CD27 in human cell culture supernatant samples.

Interpolated concentration of native CD27 was measured in duplicate at different sample concentrations and interpolated from the CD27 standard curves. Undiluted samples were 100% cell supernatant. The interpolated dilution factor corrected values were plotted (mean +/- SD, n=2). The mean CD27 concentration was determined to be 3041 pg/mL in Daudi and 3777 pg/ml in Raji in cell culture supernatant, undetectable in MCF7 cell culture supernatant.

Interpolated concentrations of spiked CD27 in cell culture media samples.

The concentrations of CD27 were measured in duplicates, interpolated from the CD27 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).

Interpolated concentrations of spiked CD27 in cell culture media samples.

The concentrations of CD27 were measured in duplicates, interpolated from the CD27 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).