qPCR Efficiency Calculator
Calculate amplification efficiency from a qPCR standard-curve slope, then interpret whether the assay is inside the practical 90-110% range.
Slope to qPCR efficiency
10-fold Ct spacing at 100% efficiency is 3.322 cycles.
Interpretation
Calculated result
qPCR efficiency from standard curve Slope: -3.35 Amplification factor: 1.988 Efficiency: 98.84% Intercept: 38.2 Replicates: 3 Expected Ct spacing for 10× dilution: 3.35 cycles Interpretation: 90–110% is a common reference range, not proof of assay validity. Also inspect R², residuals, melt curve, and assay-specific validation.
qPCR Standard Curve Linear Regression & Slope QC Chart
The x-axis shows the log10 template dilution and the y-axis shows quantification cycle (Ct). With a 10-fold dilution series and ideal amplification, 100% efficiency corresponds to a slope of about −3.322 and a Ct difference of about 3.32 cycles per dilution. A deviation is a reason to review assay conditions, not a diagnosis by itself.
qPCR Amplification Efficiency Diagnostic Matrix
| Slope Range | Efficiency (E) | Amp Factor | QC Verdict | Root Causes & Action Plan |
|---|---|---|---|---|
| < −3.58 | < 90% | < 1.90 | Low Efficiency | Annealing temperature too high; strong secondary structure; suboptimal primer design; reagents or dNTP degraded. |
| −3.58 ~ −3.10 | 90% ~ 110% | 1.90 ~ 2.10 | Common reference range | A value from 90–110% is a common reference range, not proof of MIQE compliance. Also review R², residuals, melt curves, and assay-specific validation. |
| > −3.10 | > 110% | > 2.10 | Apparent over-efficiency | Co-purified inhibitors in stock template diluting out in lower points, or non-specific primer-dimers artificially elevating signal in high dilutions. |
| Any Slope | Correlation | R² < 0.985 | Non-linear | Severe serial dilution pipetting inaccuracy, unmixed tubes, bubbles in optical read path, or evaporation from plate edges. |
Worked example
If the standard curve slope is -3.35:
How to interpret the result
90-110% is commonly acceptable for qPCR standard curves.
Below 90% suggests poor amplification, inhibitors, primer issues, or dilution mistakes.
Above 110% is often a warning sign for contamination, non-specific signal, or non-linear curve points.
Why the standard curve quality matters
The slope is only as good as the dilution series behind it. Use at least five dilution points, keep the dilution factor consistent, and inspect whether low-template points are still linear before trusting the efficiency.
Related next steps
Need to prepare the dilution series after deciding copy number targets?
Assay check
This page is scoped to slope-based qPCR efficiency and standard-curve interpretation. Use a separate plate map when you need to document well positions, replicates, and controls.
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