Dilutions of DMSO were prepared in assay buffer to give a range of concentrations from 2% (v/v) to 25% (v/v) in the wells. From the results shown in figure 2 it can be seen that the assay was tolerant to DMSO up to a final concentration of 12.5% (v/v)
Competitive binding of 0.37-nM [125I]GRO-αwith unlabeled IL-8 and GRO-αwas assessed and the IC50 values calculated as shown in Figure 3. DMSO was included at an assay concentration of 2.5% (v/v). Final concentrations in the well for unlabeled GRO-αwere 0.03 to 440 nM, and for unlabeled IL-8, 0.03 to 500 nM.
The IC50 value for GRO-αwas determined to be 2 nM (95% confidence interval range 1.7 to 2.3 nM) and the Ki value was 1.02 nM (95% confidence intervals 0.87 to 1.19 nM). The IC50 value for IL-8 was 11 nM (95% confidence interval range 10 to 12 nM) and the Ki was 5.68 nM (95% confidence intervals 5.14 to 6.27 nM).
A time course was performed using standard reagent concentrations as detailed in the protocol and DMSO included at an assay concentration of 2.5% (v/v). Readings were taken over an increased incubation time of 18 h. The assay reached equil