ΔΔCt Calculator

How to calculate fold change with ΔΔCt

1. Type the Ct values of the target gene and the reference gene for the treated and control samples. Several replicates separated by spaces are averaged.
2. Leave both efficiencies at 100% for the 2^-ΔΔCt method, or enter the measured efficiencies to use the Pfaffl method.
3. The fold change of the treated sample over the control appears, with ΔCt, ΔΔCt and the log2 fold change.

Frequently asked questions

What is the formula?

ΔCt = Ct target − Ct reference for each sample, ΔΔCt = ΔCt treated − ΔCt control, and fold change = 2^-ΔΔCt.

When should I use primer efficiencies?

When they differ from 100% or from each other. The Pfaffl ratio is E target^(Ct target control − Ct target treated) ÷ E reference^(Ct reference control − Ct reference treated), with E = 1 + efficiency. At 100% for both it gives the same result as 2^-ΔΔCt.

What does a fold change below 1 mean?

The target is expressed less in the treated sample than in the control. A fold change of 0.5 is a log2 fold change of −1, half the expression.