How to use (3 steps)
- Select an example or paste standards (concentration and signal).
- Choose 4PL/5PL, weighting, and other settings.
- The curve and unknown concentrations appear (extrapolation is flagged).
This is a model. Extrapolation outside the standard range can be inaccurate. Use blank subtraction only when needed.
Inputs (standards, unknowns, settings)
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Display options
Minimum columns: concentration and signal. Multiple rows with the same concentration are treated as replicates.
Minimum columns: sample and signal. If dilution_factor is blank, it is treated as 1.
Results (curve & concentrations)
R² and AIC are guides. Also check residuals and whether values are extrapolated.
Standard curve
Residual plot (optional)
Parameters (A, B, C, D, G)
| param | value | 95% CI (approx) |
|---|
Guide: A=Bottom (low concentration), D=Top (high), C=EC50 (midpoint), B=Hill slope, G=asymmetry (5PL only). Internally we compute (x/C)^B as exp(B·ln(x/C)). Log10 on x-axis is for readability.
QC (back-calculation / recovery)
| label | conc_nominal | signal_mean | conc_hat | recovery |
|---|
Unknowns (y → concentration)
| sample | signal_mean | DF | conc_measured | conc_original | flag |
|---|
How it’s calculated
- Fit a 4PL/5PL logistic curve with nonlinear least squares.
- Unknown concentrations are computed by inverting the fitted curve.
- Weighting (1/y, 1/y², etc.) can help handle variance across concentration ranges.
Values are guides. Also check outliers, standard ranges, and dilution conditions.
FAQ
Should I use 4PL or 5PL?
4PL is often sufficient. Use 5PL if asymmetry is needed (compare with AIC, etc.).
Do I need weighting (1/y, 1/y²)?
Not required. Start without weighting and try it if bias appears across the range.
How should I handle concentration 0 (blank)?
By default, blanks are excluded from fitting (works well with log x). Use blank subtraction only if needed.
An unknown value is outside the standard range.
It will be shown as extrapolated. Accuracy drops, so dilute and remeasure within range.
Related tools
- qPCR standard curve calculator
Similar quantification workflow (standards → curve → unknowns).
- A260 concentration & purity
Helpful for sample prep and QC.
- DNA/RNA concentration ↔ mol ↔ copy number
Another quantification tool for nucleic acids.
Comments
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