Purpose: Import pClamp files and perform common types of data analysis, mainly of biophysical properties of voltage-gated ion channels studied with voltage-step or voltage-ramp protocols, such as construction of current-voltage relationships, activation and inactivation curves, and fitting them by Boltzmann function to obtain the various parameters of various voltage-gated ion conductances. In addition, several tools for facilitation of data graphing and presentation (mask/unmask capacitive artifacts, leak correction, and data smoothing) are included in this application. Possible variations in the user voltage protocols are extensively addressed via user interface.
Description of button functionality:
1. pClamp data import. Normally, one file is imported for subsequent data analysis by pressing this button. However, multiple files can also be selected, in which case pressing the button repeatedly opens the subsequent pClamp files (e.g. file viewer function). Data analysis can be commenced once the correct file was located.
2. Data smoothing by N number of points: each point is replaced by the mean of N/2 proceeding and N/2 subsequent points.
3. Masks capacitative artifacts associated with voltage steps and automatically rescales current traces (this procedure will have to be acknowledged in Methods).
4. Undo data points masking.
5. Current-voltage (I-V) relationship with options to measure max, min or mean values, or any combinations of min/max/mean values in different episodes (these options are more versatile than in Clampfit).
6. Input resistance calculation from the I-V curve and leak current correction (e.g. subtraction of a linear component of the I-V curve).
7. Activation (conductance) curve is created and automatically fitted by the Boltzmann function with the preset lower bound Gmin≥0 (since conductance should be ≥0) to determine the potential of half-maximal activation (V1/2) and slope factor (k).
8. Inactivation curve is created and automatically fitted by the Boltzmann function to determine the potential of half-maximal inactivation (V1/2) and slope factor (k).
9. Extracts I-V relationships from voltage ramp protocols. If file contains multiple episodes with the same voltage ramp protocol then all I-V curves are extracted in one go (this option is not available in Clampfit). If voltage protocol used contains several ramp epochs a pop-up dialog allows the user to choose any one for constructing the I-V curves.
10. Calculation of conductance-voltage (G-V) relationship based on ramp I-V curve: reversal potentials of the ramp I-V curves are measured by linear regression of data points around the estimated reversal potential to increase the accuracy, and based on these values the conductance curves are created with the option of simultaneously fitting them by the Boltzmann function. Slope factor can be shared for the cases when the activation curves shift on the voltage axis in a parallel manner. This tool is particularly suitable for analysis of cationic currents.
11. Quick Tutorial is provided in the form of PowerPoint slides with all steps of data analysis described in the Notes for each slide. Sample files used are included allowing the user to learn how to use this App for various types of pClamp data analysis. Sample files are saved in Apps folder: \\Simple pCLAMP Analyzer\Samples. Type "%@A=" in Script window to see App folder.