qPCR slope to efficiency

Convert the slope of a qPCR standard curve into amplification efficiency. Enter the slope below to get the efficiency as a percentage and the fold amplification per cycle, with a quick quality check.

How efficiency relates to the slope

You build a standard curve by plotting quantification cycle (Cq/Ct) against the log of template dilution; its slope gives the efficiency:

Efficiency = 10(−1/slope) − 1

A perfect reaction doubles the product every cycle, which corresponds to a slope of −3.32 and 100% efficiency. In practice, efficiencies of 90–110% (slopes of about −3.6 to −3.1) are considered acceptable.

Getting a reliable slope

• Run a dilution series (e.g. 5–6 points, 10-fold steps) of a known template, in replicates.
• Plot Cq vs log(dilution) and fit a line; the software reports the slope and R².
• Aim for R² > 0.98 — a poor fit makes the efficiency unreliable.

Troubleshooting

• Efficiency < 90% (slope steeper than −3.6): inhibitors in the template, suboptimal primers, or pipetting error in the dilutions.
• Efficiency > 110% (slope shallower than −3.1): often pipetting inaccuracy at high dilutions, primer-dimers, or non-specific product.
• For the ΔΔCq method, target and reference assays should have similar, near-100% efficiencies; otherwise use an efficiency-corrected model.

Frequently asked questions

What slope means 100% efficiency?

A slope of −3.32, which corresponds to the template doubling every cycle.

What is an acceptable qPCR efficiency?

Generally 90–110% (slopes of about −3.6 to −3.1), with R² > 0.98.

How do I calculate efficiency from the slope?

Efficiency = 10^(−1/slope) − 1, expressed as a percentage. Enter your slope above to compute it.

Why is my efficiency above 100%?

Usually pipetting inaccuracy at high dilutions, primer-dimers or non-specific amplification — check the curve and melt profile.

Related guide: qPCR data analysis →