2-Dimensional Spectrum Analysis:

The 2-dimensional spectrum analysis is used for composition analysis of sedimentation velocity experiments. It can generate sedimentation coefficient, diffusion coefficient, frictional coefficient, f/f0 ratio and molecular weight distributions. The distributions can be plotted as 3-dimensional plots (2 parameters from the above list against each other), with the 3rd dimension representing the concentration of the solute found in the composition analysis.

You can start the 2-dimensional spectrum analysis in one of two ways: 1. by clicking on "2-D Spectrum Analysis" inside the Enhanced van Holde - Weischet method (the preferred approach), or 2. by clicking on "2-D Spectrum Analysis" in the Velocity sub-menu of the main menu. If you started the analysis routine from the enhanced van Holde - Weischet analysis windows, the main data analysis window will appear with the same dataset loaded that was analyzed by the van Holde - Weischet analysis, and most parameters will have been appropriately initialized.

If you started the analysis routine from the main menu, the main data analysis window will appear without any data loaded. In that case, you will need to load an UltraScan dataset that has previously been edited with the UltraScan editing module. UltraScan sedimentation velocity datasets always have the suffix ".us.v". Click on the "Load Dataset" button in the left upper corner of the control panel. Simply select the desired run from the selection of available runs in the dataset loading dialog. Once loaded, the Run Details will be shown. Click on the "Accept" or "Cancel" button, and you will be returned to the analysis window, which will now show the available dataset of the selected run in the edited data window on the lower right panel. analysis window.

Analysis Functions:

Click on these buttons to control the 2-dimensional spectrum analysis:
  • Load Data: Load edited data sets (with the *.us suffix). A file dialogue will allow you to select a previously edited and saved velocity experiment. If the data was edited with an older version of UltraScan than the current, an error message will be displayed.
  • Run Details: View the diagnostic details for a particular run.
  • Reset Data use this button to reset the default settings for the analysis.
  • Save Data: Write out a copy of all results to an ASCII formatted data file suitable for import into a spreadsheet plotting program. See "File Structures and Formats" for details.
    Note: These files are overwritten each time this button is clicked. Only the last version of the analysis will be saved!
  • Print Data: Load the printer control panel for printing of plot graphics
  • View Data Report: See the data report for the last analysis setting. Note: This file is re-written each time it is accessed. Only the current analysis result is available.
  • Help: This help file
  • Close: Close the 2-dimensional spectrum analysis window.

Run Information:

  • Run ID: The name of the run given during editing
  • Temperature: The average temperature calculated from the entire run
  • Available Cells: The numbers the cells that contain analyzable data
Clicking on a cell and wavelength selection will bring up the cell contents description for that cell and wavelength. Scroll through this list to bring up information for cells > 3. If there is no data available for the selected cell, the program will list "No Data available". Selecting a cell/wavelength combination will automatically bring up the corresponding dataset and present the analysis.

Experimental Parameters:
Here you can enter the corrections for density, viscosity, and partial specific volume of your sample. As you change the information in these fields, the program will automatically update the analysis to correct the S-value according to the specified buffer conditions. If the data was retrieved from the database and associated with the correct buffer file and peptide file, the density, viscosity and partial specific volume (vbar) will automatically be updated to the appropriate values and don't need to be adjusted to produce S 20,W corrected values.
  • Density: Click to calculate the density based on the composition of your buffer.
  • Viscosity: Click to calculate the viscosity based on the composition of your buffer.
  • vbar(20o): Click to calculate the partial specific volume for a peptide based on its primary amino acid sequence and correct the vbar value for the current temperature.
  • Fit Control: Click here to open the fitting control window.

Analysis Controls:

  1. Status Info: This field contains information about the fitting progress.
  2. Exlude Single Scan: When setting this counter to a non-zero value, the respective scan will be highlighted in red. Clicking on "Excl. Single Scan" while a scan is highlighted in red will delete this scan from the analysis. Deleting scans from the analysis is irreversible and can only be reset by clicking on the "Reset" button or by reloading the data (when smoothing, the data is always automatically reloaded, causing the number of scans to be reset to the original count). Skipped scans are excluded automatically, because they haven't cleared the meniscus. This is equivalent to scans whose first point has an absorbance larger than the baseline (as defined by the baseline absorbance selected during editing). Second moment points for these scans will be displayed in red and excluded from the average, which is displayed as a horizontal line in the second moment plot.
  3. Exclude Scan Range: Same as "Exclude Single Scan", except for multiple scans. To use this feature, select first the start scan of the range by using "Exclude Single Scan", then complete the scan range by using "Exclude Scan Range". The fewer scans are included, the greater the effect of noise on the fit, but the fewer megabytes of memory are used during fitting.
  4. Status: The status bar informs the user about the progress of the analysis. On slower computers this has more significance than on a fast Unix machine.

Utility of the 2-dimensional spectrum analysis:

This method decomposes the velocity experimental data into a sum of non-interacting finite element solutions, which provide information on sedimentation and shape. A publication describing this method in more detail is in the works. More information about this method will be provided in a future release.

www contact: Borries Demeler

This document is part of the UltraScan Software Documentation distribution.
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Last modified on July 17, 2005.