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Melt curve analysis

Willem Geerts
Updated

    Another acquisition type is qPCR melt curve analysis, which is supported by FastFinder Analysis. The setup for a melt-based analysis is very similar to the standard configuration and lay-out of FastFinder Analysis (i.e. amplification as acquisition type). The main differences between an analysis for amplification versus melt will be highlighted here.

    PCR setup

    There are no specific features or settings for melt. For more information on the PCR setup tab, please consult Analysis - PCR setup.

    Summary

    In the Summary tab, the Control and Sample results are provided exactly as for a regular amplification analysis, with the exception that for the individual targets the melt temperature (Tm) is calculated (instead of a Cq value).

    Melt Summary_sample results.png

    Details

    In the Details tab, an overview of the individual target result is provided. This includes the result state and if detected, the melt temperature.

    Melt Details_Target results.png

    Graphical layout

    In the graph, the melt curves are visualized for each of the sample targets.

    Symbol

    Meaning
    mceclip0.png

    Move graph area
    mceclip1.png

    Zoom to area
    mceclip2.png

    Lasso selection
    mceclip3.png

    Reset graph zoom
    mceclip4.png

    Automatic graph zoom
    mceclip5.png

    Show full screen
    mceclip7.png

    Graph settings

     

    Under the graph settings, the graph features and lay-out can be adjusted:

    • The curve color can be organized either by dye or by result.
    • The chart type can be set to melt curve or melt peaks:
      • Melt peak: visualization of the data following processing by the algorithm (y-axis = -d(RFU)/dT).
      • Melt curve: visualization of the raw melt data (y-axis = RFU).
    • Enable or disable the option to show only your selected data (other data is removed from the background) or all data.
    • Enable showing Tm values for the selected data.
    • Permanently show control curve data when selecting a sample.
    • Fill in your preferred X- and Y- axis min/max values.

    Press "Save" to save your changes or "Reset scales" to go back to the default settings.

    Melt Graph.png

    Plate layout

    Similar to amplification analyses, the plate layout shows the results on target or assay target level and allows the user to navigate the results in a more visual manner if desired.

     

     

     

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