Mouse Osteopontin Recombinant Rabbit Monoclonal Antibody [PSH17-50] - BSA and Azide free (Detector)

Sandwich ELISA analysis of Mouse Osteopontin matched pair antibodies

Capture: HA723924, Mouse Osteopontin Rabbit mAb [PSH17-49]
Detector: HA723925, Mouse Osteopontin Rabbit mAb [PSH17-50]

Elisa assay was performed by coating wells of a 96-well plate with 100 µl per well of capture antibody (HA723924) 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 Mouse Osteopontin protein (HA210997) protein starting from 2,500 pg/ml to 0 pg/ml and detect antibody (HA723925, 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 spp1 in RAW 264.7 cell extract samples based on a 1000 µg/ml extract load.

Capture: HA723924, Mouse Osteopontin Rabbit mAb [PSH17-49]
Detector: HA723925, Mouse Osteopontin Rabbit mAb [PSH17-50]

Interpolated concentration of native spp1 was measured in duplicate at different sample concentrations. The interpolated dilution factor corrected values were plotted (mean +/- SD, n=2). The mean spp1 concentration was determined to be 79,438,854 pg/ml in RAW 264.7 treated cell extract and 6,041,158 pg/ml in RAW 264.7 untreated cell extract.

Interpolated concentrations of native spp1 in Mouse serum.

Capture: HA723924, Mouse Osteopontin Rabbit mAb [PSH17-49]
Detector: HA723925, Mouse Osteopontin Rabbit mAb [PSH17-50]

Interpolated concentration of native spp1 was measured in duplicate at different sample concentrations and interpolated from the spp1 standard curves. The interpolated dilution factor corrected values were plotted (mean +/- SD, n=2). The mean spp1 concentration was determined to be 77,206 pg/ml in Mouse serum.

Interpolated concentrations of native spp1 in J774A.1 cell culture supernatant.

Capture: HA723924, Mouse Osteopontin Rabbit mAb [PSH17-49]
Detector: HA723925, Mouse Osteopontin Rabbit mAb [PSH17-50]

Interpolated concentration of native spp1 was measured in duplicate at different sample concentrations and interpolated from the spp1 standard curves. The interpolated dilution factor corrected values were plotted (mean +/- SD, n=2). The mean spp1 concentration was determined to be 92,201 pg/ml in J774A.1 cell culture supernatant.

Interpolated concentrations of spiked spp1 in cell culture media samples.

Capture: HA723924, Mouse Osteopontin Rabbit mAb [PSH17-49]
Detector: HA723925, Mouse Osteopontin Rabbit mAb [PSH17-50]

The concentrations of spp1 were measured in duplicates, interpolated from the spp1 standard curves and corrected for sample dilution. diluted samples are as follows: 50% cell culture media with FBS. The interpolated dilution factor corrected values are plotted (mean +/- SD, n=2).