Rat IFN gamma Recombinant Rabbit Monoclonal Antibody [PSH05-69] - BSA and Azide free (Capture)

Sandwich ELISA analysis of rat IFNG matched pair antibodies

Elisa assay was performed by coating wells of a 96-well plate with 100 µl per well of capture antibody (HA722421) diluted in carbonate/bicarbonate buffer, at a concentration of 5 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 standard Rat IFN gamma protein (HA210577) starting from 2,500 pg/ml to 0 pg/ml and detect antibody (HA722422, 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 IFNG in Concanavalin A treated rat spleen(18 hours) supernatant samples.

Rat spleen was cultured for 18 hours in the presence or absence of 3 µg/ml Concanavalin A. The concentrations of IFNG measured in duplicate and interpolated from the IFNG standard curve and corrected for sample dilution. Undiluted samples are as follows: unstimulated 6% and stimulated 6%. The interpolated dilution factor corrected values are plotted (mean +/- SD, n=2). The mean IFNG concentration was determined to be 10.8 ng/ml in Concanavalin A stimulated rat spleen supernatant and undetectable in unstimulated rat spleen supernatant.

Interpolated concentrations of spiked IFNG in rat cell culture media samples.

The concentrations of IFNG were interpolated from the IFNG standard curves and corrected for sample dilution. Undiluted samples are as follows: cell culture media 50%.