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Sandwich ELISA analysis of human MMP-1 matched pair antibodies
Elisa assay was performed by coating wells of a 96-well plate with 100 µl per well of capture antibody (HA722874) 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 CXCL13 protein (HA210903) starting from 3000 pg/ml to 0 pg/ml and detect antibody (HA722875, 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.
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Interpolated concentrations of native Pro-MMP-1 in human cell culture supernatant samples.
Interpolated concentration of native Pro-MMP-1 was measured in duplicate at different sample concentrations and interpolated from the Pro-MMP-1 standard curves. Undiluted samples were 100% cell supernatant. The interpolated dilution factor corrected values were plotted (mean +/- SD, n=2). The mean Pro-MMP-1 concentration was determined to be 2316 pg/mL in SW480 and 1660 pg/ml in HepG2 in cell culture supernatant, undetectable in A549 supernatant.
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Interpolated concentrations of spiked Pro-MMP-1 in human cell culture media samples.
The concentrations of Pro-MMP-1 were measured in duplicates, interpolated from the Pro-MMP-1 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).
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