1. What is a qPCR Efficiency Calculator?

Definition: This calculator determines the amplification efficiency of a qPCR reaction based on the slope of the standard curve.

Purpose: It helps molecular biologists assess the performance of their qPCR assays, which is crucial for accurate quantification.

2. How Does the Calculator Work?

The calculator uses the formula:

Where:

• \( Eff \) — PCR efficiency (decimal)
• \( slope \) — Slope of the standard curve (dimensionless)

Explanation: The slope of the standard curve is determined from the plot of Ct values versus the log of template concentration. The efficiency is then calculated from this slope.

3. Importance of qPCR Efficiency

Details: Ideal qPCR efficiency is 100% (Eff = 1), meaning the amount of product doubles each cycle. Efficiency between 90-110% is generally acceptable.

4. Using the Calculator

Tips: Enter the slope of your standard curve (typically between -3.1 to -3.6 for good efficiency). The calculator will provide both decimal and percentage efficiency values.

5. Frequently Asked Questions (FAQ)

Q1: What is an ideal slope value?
A: The ideal slope is -3.32, which corresponds to 100% efficiency. This means the PCR product doubles exactly each cycle.

Q2: What slope range is acceptable?
A: Slopes between -3.1 to -3.6 typically indicate efficiencies between 90-110%, which are generally acceptable.

Q3: What does low efficiency indicate?
A: Low efficiency (<90%) may indicate PCR inhibition, poor primer design, or suboptimal reaction conditions.

Q4: What does high efficiency indicate?
A: Efficiency >110% may indicate pipetting errors, contamination, or non-specific amplification.

Q5: How do I determine the slope?
A: The slope is obtained from the standard curve plot generated by your qPCR software when using serial dilutions of a known template.