Melt Curve Peak Tm Identifier Calculator

Calculator Input

Paste two-column data in any common format. Commas, tabs, spaces, and semicolons are accepted. The first column is temperature, and the second is fluorescence.

Sample name

Assay note

Temperature unit

Smoothing window

Odd values work best for moving-average smoothing.

Minimum peak height

Minimum prominence

Minimum peak spacing

Spacing is measured in the chosen temperature unit.

Maximum peaks to report

Apply baseline correction to smoothed fluorescence

Temperature and fluorescence pairs

Header rows are allowed. Example: 78.0, 642.15

Reset

Example Data Table

This sample shows the first ten rows from the built-in demonstration dataset.

Temperature (°C)	Fluorescence
68.00	981.15
69.00	983.43
70.00	984.97
71.00	985.41
72.00	984.63
73.00	982.66
74.00	979.65
75.00	975.58
76.00	969.78
77.00	959.88

Formula Used

1) Moving-average smoothing
F̄_i = (Σ F_j) / n across the selected window.

2) Negative first derivative
-dF/dT ≈ -[(F̄_i+1 - F̄_i-1) / (T_i+1 - T_i-1)].

3) Peak Tm selection
Tm = temperature at the local maximum of the -dF/dT curve.

4) Peak prominence
Prominence = Peak Height - Reference Base, where the base is the larger of the left and right minima.

5) Width at half height
Width = Right crossing temperature - Left crossing temperature at half-prominence level.

6) Confidence score
Score = 50×height term + 30×prominence term + 20×width term, capped at 100.

This calculator identifies candidate melt peaks from fluorescence-versus-temperature data. It smooths the signal, optionally baseline-corrects it, calculates the negative derivative, and ranks local maxima by strength and separation.

How to Use This Calculator

Paste temperature and fluorescence pairs into the data box.
Choose your temperature unit and add a sample name.
Set smoothing, minimum peak height, prominence, and spacing.
Turn baseline correction on when background offset affects raw intensity.
Click Identify Peak Tm to calculate the derivative and detect peaks.
Review the primary Tm, confidence score, and all reported peaks.
Use the Plotly chart to compare fluorescence and derivative behavior.
Download the result summary as CSV or PDF for reporting.

Frequently Asked Questions

1) What is a melt curve peak Tm?

It is the temperature where the melt transition produces the highest negative derivative signal. In practice, that temperature often represents the main duplex dissociation point for the amplified product.

2) Why does the calculator use -dF/dT?

Melt analysis commonly transforms fluorescence decline into a derivative curve. The local maximum of the negative derivative makes the melt transition easier to locate than inspecting raw fluorescence alone.

3) What do multiple peaks usually mean?

Multiple peaks can suggest non-specific amplification, primer-dimers, mixed amplicons, or heteroduplex formation. Context still matters, so compare results with assay design, controls, and instrument output.

4) How many data points should I enter?

At least five unique points are required, but more points usually give more stable derivatives and clearer peak detection. Dense temperature sampling improves Tm precision and peak width estimates.

5) When should I use baseline correction?

Use it when the overall fluorescence baseline shifts upward or downward and makes the derivative harder to interpret. Baseline correction can clarify peak structure without changing temperature order.

6) Can this replace instrument software?

No. It is best used for quick analysis, teaching, screening, or independent review. Final biological interpretation should still consider instrument settings, controls, dye chemistry, and validation workflow.

7) What does a broad peak indicate?

A broad peak may reflect lower resolution, mixed products, noisy data, or gradual melting behavior. Wide peaks often reduce confidence because the melt transition is less sharply defined.

8) Does the temperature unit change the math?

The algorithm uses whatever unit your data already follows. Keep all rows in one unit only. Peak spacing and width thresholds should match the same scale for consistent results.