| Assay Method Information | |
| | Antitrypanosomal Activity Assay |
| Description: | Antitrypanosomal activity was tested on T. brucei brucei bloodstream from parasites (strain Lister 427, VATMITat 1.2). Stock solutions of compounds (50 mM) were prepared in DMSO. Two fold serial dilutions of each compound (10 wells in each row) were prepared in 96 well plates in HMI111 medium (100 μl/well; quadruplicates; cf. also Hirumi et al., J. Parasitol. 75 (1989), 985-989). One well without compound, and one well with media alone were included in each row. Parasites grown in HMI11 were inoculated (100 μl of 4×104/ml) in all wells (except the well with media alone ) giving a final concentration of parasites of 2×104/ml. The plates were incubated for 66 hours at 37° C. in the incubator with 5% CO2. 20 μl resazurin (Sigma, 0.125 mg/ml in Hanks Balanced Salt Solution) was added to all wells and the plate was further incubated for 6 hours. Change in color of resazurin (blue to pink) by living cells was measured with a 96 well plate reader (excitation at 530 nm, emission at 585 nm, filter cut-off: 570 nm). Percent inhibition values were calculated by setting the well without compound to 0% inhibition . Non-linear regression graphs were plotted in Graphpad Prism to yield sigmoidal dose-response curves and IC50 values were determined (compound concentration giving 50% inhibition). |
| Affinity data for this assay | |
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