Dose response assays are generally characterized by a nonlinear relationship between the response and the amount of drug/agonist/antagonist given. When plotted, this response will usually exhibit a sigmoidal curve that is best described using the 4 Parameter Logistic (4-PL) or 5 Parameter Logistic (5-PL) nonlinear regression model equations.
Dose Response Curve
As described in our Bioassay Analysis with the Five Parameter Logistic (5-PL) Non-Linear Regression Curve-Fitting Model blog post, the C parameter in both the 4-PL and 5-PL models represent the concentration value for the inflection point of the curve. Depending on whether you have an inhibition or activation assay, the C parameter also represents the half maximal inhibitory concentration (IC50) or the half maximal effective concentration (EC50) respectively.
Anatomy of the 4 or 5 Parameter Logistic Model
MasterPlex ReaderFit offers full support for the 4 Parameter Logistic (4-PL) and 5 Parameter Logistic (5-PL) model equations making it an ideal tool for automated fitting of dose response curves and calculating the EC50 or IC50 values.
Here is a Quick Guide for this procedure:
- To acquire MasterPlex ReaderFit, download and install the free 14-day trial.
- If you have not yet imported your data, please refer to the community post on importing data into MasterPlex ReaderFit.
- This is a sample dose response data set which contains 3 different sets of data from which we will generate 3 dose response curves (each outlined with a different color). Each curve set contains 8 data points with duplicates that are side-by-side.
Sample Dose Response Data
- Select 1 entire set of data points (including replicates) and click the blue Standard button. Please note that you can hold down the CTRL button and left click non-contiguous wells.
Marking Data Sets
- The Auto Fill dialog will pop up. This feature is only helpful if you have a serial dilution for your drug concentrations. If not, you will have to enter them in manually.
Auto Fill Feature for Entering in Serial Dilution Concentrations
In this example, Drug ABC was serially diluted from top (20,000 pg/mL) to bottom (0.26 pg/mL) with a dilution factor of 5.
- Click Apply & View to preview the values in plate tab to confirm.
- Repeat steps 2-4 for any other data sets you may have. In this example, there are 2 other data sets.
For each cell, you will notice 2 numbers: the response value on the top and the drug dose on the bottom. - Click on the Standard Curve tab.
- Make sure the Use Best Fit feature checkbox is unchecked and select the model equation (4-PL or 5-PL) you would like to use for the curve fit and then click Calculate.
Model Equation Selection and Calculation
- MasterPlex ReaderFit will then draw the dose response curve(s) according to the parameters you have chosen.
Dose Response Curves based on the 4 Parameter Logistic
- To view the EC50 or IC50 calculations, click on the individual curves (if you have more than one) and look for the C parameter listed above the dose response curve. In this case, this value represents the EC50 because this was an activation assay.
EC50 value
For more information or to download the free 14-day trial, please visit the MasterPlex ReaderFit product page.
