What the IC50 is
The IC50 (half-maximal inhibitory concentration) is the concentration of a drug or inhibitor that reduces a response, such as enzyme activity or cell viability, halfway between its top and bottom levels. For a response that rises with concentration, the same point is called the EC50.
Designing the experiment
- Test 8 to 12 concentrations spaced evenly on a log scale, such as threefold or tenfold steps, so the curve has points on both plateaus and through the middle.
- Include an untreated control (100%) and a full-inhibition or background control (0%).
- Run each concentration in replicate.
Normalizing the data
Convert each reading to a percentage of the controls:
% response = (reading − background) ÷ (untreated − background) × 100
Fitting the curve
Dose-response data form an S-shaped curve against log concentration, fitted with the four-parameter logistic model:
response = bottom + (top − bottom) ÷ (1 + 10(log IC50 − log concentration) × Hill slope)
The fit finds the top and bottom plateaus, the IC50 at the midpoint, and the Hill slope, which describes the steepness. If your data do not reach a plateau, fixing top at 100 and bottom at 0 after normalizing gives a more stable fit.
Reading the result
- IC50: a lower value means a more potent compound.
- Hill slope: about −1 for a simple falling curve. Much steeper or shallower slopes are worth a second look.
- R² and the plot: the curve should run through the points without a systematic pattern of misses.
- An IC50 outside the tested range is an extrapolation and should be confirmed with more concentrations.
Calculators
The IC50 Calculator fits the four-parameter curve to your replicates, draws it and gives the IC50 or EC50, Hill slope, plateaus and R². Plan the concentrations with the Serial Dilution Calculator.