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Calculating the EC10 or IC90 values for a dose response curve is not as simple as the EC50 or IC50 values when using the 4PL or 5PL model equations.

The equation for calculating any EC value given a 4PL curve fit is the following:

EC_F = (F/(100-F))^(1/H) * EC₅₀

0 < F < 100 (This is the EC value you are interested in. For example, F=10 for EC₁₀.)
H = Hill Slope for the 4PL curve fit

Luckily, our latest release of MasterPlex ReaderFit 2010 makes this easy. Yes, even with the 5PL curve-fit!

Here’s how:

1. Copy & paste your data into ReaderFit

   Copy your response values from the source and paste it into ReaderFit via CTRL+V -> Paste Response Values.

2. Mark data as Sample Curve type (part 1)

   Select all of the data points that will be used to fit one entire dose response curve including all replicates. In the example given below, there are 7 groups each done in duplicate.
   

3. Mark data as Sample Curve type (part 2)

   Click on the blue Mark Sample Curve button. (The Auto Fill dialog may pop up. Click the Close button if you see it.)
   

4. Indicate known dose concentration values (part 1)

   Select Independent Values(X-axis) from the data pull-down menu. This will show the corresponding data in the plate view. By default, all of the Independent Values(X-axis) will be 0.
   

5. Indicate known dose concentration values (part 2)

   Select the sample of interest and double-click the lower-cell to enter edit mode. Once in edit mode, type in the concentration value. Make sure all values have the same unit. We will edit that later.
   

6. Fitting the curve (part 1)

   Select the Fit Curves tab.
   

7. Fitting the curve (part 2)

   Select either the Four Parameter Logistic (4PL) or Five Parameter Logistic (5PL) from the Curve Fitting section and press the Calculate button.
   

8. Edit dose response curve axes

   Right-click on the dose response curve chart and edit the x and y axes. You also have the option to add a chart title.
   

9. Calculate EC or IC(Anything) value (part 1)

   Press the Statistics Toolbar button.

10. Calculate EC or IC(Anything) value (part 2)

    By default the EC50 and the log EC50 values are calculated for you. If you have an inhibition curve, press the ICxx radio button to switch to IC50 and log IC50 values.
    

11. Calculate EC or IC(Anything) value (part 3)

    If you have an inhibition curve and you are interested in the IC90 calculation, simply select the ICxx radio button and enter “90” into the Percentages section and press the Add button.
    

If you have not done so yet, I would like to invite you to try the free 14-day trial of MasterPlex ReaderFit.

We recently had a customer that was interested in MasterPlex QT because his current analysis software for his Bio-Plex instrument was reporting a lot of “OOR < ” or out of range concentration values (below the lower asymptote in this case) for points on the lower end of curve. This is what you would normally expect to see for values that fall below the minimum asymptote BUT the software did not have the capability to use weighting in calculating the lower asymptote which can greatly affect points on the lower part of the curve.
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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 equation.

Dose Response Curve

What’s New in this Upgrade?

• For MasterPlex QT
• For MasterPlex ReaderFit
• For MasterPlex EX

The following webinar recordings have been made available for viewing at your own leisure:

• Reduce cost and length by 50% or more on whole-genome sequencing projects
  View Webinar | PowerPoint Presentation
• ELISA Curve-Fitting Analysis with ReaderFit.com
  View Webinar | PowerPoint Presentation
• How can Optical Mapping accelerate my research?
  View Webinar | PowerPoint Presentation
• MasterPlex ReaderFit 101
  View Webinar | PowerPoint Presentation
• How to create the report you want out of MasterPlex?
  View Webinar | PowerPoint Presentation
• MasterPlex: One Quantitative Analysis Software Solution for Multiple Instruments
  View Webinar | PowerPoint Presentation
• Getting the Most Out of Your Multiplex Data
  View Webinar | PowerPoint Presentation

Join us for our first webinar on using MasterPlex ReaderFit for microplate reader or elisa data analysis.

Join us for a Live FREE Webinar on August 6 2009 11:00 AM – 12:00 PM PDT:

Microplate Reader/Elisa Data Analysis made easy with MasterPlex ReaderFit

Space is limited. Reserve your Webinar seat now at:
https://www1.gotomeeting.com/register/364896864

We will be covering the following topics in detail:

1. File Import Wizard
2. Automatic Detection of Outliers
3. Marking Wells and Grouping of Replicates
4. Entering Standard Information
5. Best Fit and Best Fit Parameters
6. Plotting Unknown Data Points on the Standard Curve
7. WYSIWYG Data Table
8. Chart Customization
9. Templates
10. Custom Reports
11. Virtual Plate

Here is a small MasterPlex Readerfit demonstration if you simply cannot wait.

We hope to see you there!

Quick Demo of MasterPlex ReaderFit (1:43 min.)

The microplate analysis demo includes the following simple steps:

1. Importing an Excel file with the RLU values
2. Marking the background, standard, and unknown samples
3. Filling in the standard concentrations with the AutoFill Feature
4. Pressing the Best Fit button
5. Reviewing standard curve, concentrations, and charts

Save time, stop copying & pasting, and download the FREE MasterPlex ReaderFit trial and see for yourself how easy it can be!

The Hitachi Software MiraiBio group has just released a new build for MasterPlex ReaderFit, the quantitative analysis and curve-fitting data analysis software for microplate readers. New features and improvements include:

• File Import Wizard – Although MasterPlex ReaderFit can automatically import some of the known file formats from common plate readers that are out there in the market, it does not handle all 100% of them. Unknown file formats can now be handled by the File Import Wizard in 1 of 2 ways:
  1. Plate Format

     

  2. List

     

  More details on using the File Import Wizard for MasterPlex ReaderFit

• Best Fit Options – The Best Fit feature now offers 3 parameters for which you can optimize for to determine the best curve fit:
  1. Root Mean Square Error (RMSE) – RMSE is a good measure of accuracy or goodness of fit which aggregates the individual differences or residuals between your expected and calculated values into a single measure of predictive power. This is the default option as it is the one that is recommended.
  2. Standard Deviation of % Recovery – This option will provide the model equation and weighting algorithm combination with the lowest standard deviation of percent recovery. The percent recovery is basically the calculated concentration – the expected concentration * 100.
  3. R-squared – This option is the only Best Fit option in previous builds of MasterPlex ReaderFit. It is not as sensitive to the goodness of fit as the RMSE option.

  

• Competitive Binding and Inhibition Assays – This build provides improved support for competitive binding or inhibition assays.

  

• More Intuitive Auto Fill – The Auto Fill feature helps to automate the entry of the standard concentration values for serial dilutions. This feature has been simplified with more intuitive dilution direction buttons and being able to preview the standard values.

  

Try it out and download the free trial now!

The 5-PL or 5 Parameter Logistic is a nonlinear regression model used for prediction of the probability of occurrence of an event by fitting data to a logistic curve. It differs from the 4-PL or 4 Parameter Logistic model in that it is an asymmetric function which is a better fit for immunoassay or bioassay data.  As the name suggests, there are 5 parameters in the 5-PL model equation:

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