Manual

Equilibrium Simulator:

You start the Equilibrium Simulator by clicking on main menu from UltraScan. The main Equilibrium Simulator Window will appear. This module can be used as an educational tool to simulate equilibrium experiments at multiple speeds and concentrations. Several predefined and user-definable models are available. All scan parameters such as the number of datapoints, maximum OD, centerpiece geometry, pathlength, can be adjusted by the user. Sample properties such as composition, extinction coefficient, vbar, molecular weight and loading concentration, as well as buffer density, baseline and noise levels can be adjusted to simulate every aspect of the experiment. Optical density scan profiles can be plotted, and a concentration histogram can be displayed. Simulated data can be exported to binary UltraScan format, or to the ASCII format used in the Beckman data acquisition program. The internal UltraScan formatted data can be used directly without further editing in the equilibrium analysis modules. Beckman formatted data needs to be edited with the UltraScan editing modules before analysis with UltraScan.

Explanation of Fields and Buttons:

Model Selection: To select a model, click on the "Select Model" button. You can then select a predefined or user-defined model from a model selection dialog. Model selection is explained in more detail here.

Component Information:

Species: For multiple component models you can use the arrow button to select the desired component.
Mol. Weight (dalton): Enter the molecular weight for the conponent selected above.
Concentration (mM): Enter the partial concentration in millimolar for the component selected above.
V-bar (ccm/g, 20^oC): Enter the partial specific volume for the component selected above. Clicking on the button will load the Vbar Calculation module, which allows you to estimate a v-bar value from the amino acid composition specified in a protein sequence file.
Ext. Coeff. (OD/cm*M): Enter the extinction coefficient of the component selected above for the the wavelength to be simulated. The units for the extinction coefficient are measured in optical density units at the selected wavelength per cm and mol. Clicking on the button will load the peptide properties calculation module, which allows you to estimate an extinction coefficient for the denatured protein at 280 nm from the amino acid composition specified in a protein sequence file.

Model Parameters:

Assoc. Const. 1/2/3: Enter the natural log of the desired association constant in molar concentration units. Fields will be disabled for models that do not require association constants.
Baseline OD: Enter the baseline concentration, if any for the simulated experiment.
Loading OD Contrib.: The figure listed in this field is the contribution to the optical density for the selected component. This number is calculated from the molar concentration and the extinction coefficient. This number cannot be changed by the user. If you want to change the value listed here, you need to adjust the molar concentration or the extinction coefficient.

Conc. Increment (OD): Enter the concentration increment for multiple loading concentrations. For a global equilibrium experiment, at least three loading concentrations should be measured (for example, 0.3, 0.5, and 0.7 OD). The increment should be listed in OD units.
# of Conc. Steps: Enter the number of desired concentration steps for this experiment. For example, if three different loading concentrations are desired click on the arrow buttons to select "3". The double arrow buttons chnage the setting in steps of 10, the single arrow button in unity steps.
Temperature (^oC): Enter the temperature in degrees Celsius for the experiment. If different from 20^oC, the program will automatically adjust the vbar and density settings to the new temperature.
Density (g/ccm 20^oC): Enter the density of the buffer for the simulated experiment. The density is entered in cgs units for the buffer at 20^oC. Clicking on the "Density" button loads the buffer calculation module.
Column Height (mm): Select the column height in millimeter for the experiment.
Meniscus: Displays the current meniscus position. The meniscus position is automatically calculated from the column height and the cell bottom position.

Pathlength (cm): Enter the centerpiece pathlength (in centimeters).
Speed Start: For a global equilibrium experiment it is recommended to use multiple speeds. The correct speed depends on the molecular weight of the component(s) to be simulated, and are best expressed in values of sigma (reduced molecular weight) ranging between 0.5 and 4. The Equilibrium Speed Estimation module can assist you in determining the best speeds for your experiment. Use this field to enter the lowest speed of the experiment. Use the arrow keys to select the speed. The single arrow key changes the speed in steps of 100 rpm, double arrow keys change the speed in steps of 1000 rpm, and triple arrow keys change the speed in units of 10,000 rpm.
Speed Stop: Select the highest speed for this simulation.
Speed Steps: Select the number of different speeds used in this simulation. The program will calculate equal speed intervals between the Speed Start value and the Speed Stop value. If only 2 speeds are selected, the Start Speed and the Stop Speed will be used. If one speed is selected, the Start Speed only will be used.
Cell Bottom (cm): Enter the cell bottom position in centimeter to define the geometry of the cell. The single arrow button changes the position in units of 1x10<^-3 cm, the double arrow button changes the position in units of 1x10<^-2 cm, and the triple button changes the position in steps of 0.1 centimeter.
Maximum OD: Enter the maximum OD to be simulated in the experiment. All data exceeding this value will not be displayed.

# of Datapoints: Select the number of datapoints for this simulation. Each scan will be simulated with that many points, which will be equally spaced wit the radial step size listed under "Radial Increment".
Gaussian Noise (% OD): Enter the percentage of the total optical density to be simulated as random noise, with gaussian distribution. Single arrow button clicks will adjust the noise level in steps of 0.01 OD percentage points, double arrow button clicks will adjust the level in steps of 0.1 OD percentage points, and single arrow button click will adjust it in steps of unity. 0.1 - 0.3 percent of 1 OD are common values for the XL-A analytical ultracentrifuge.
Nonlinear Noise (% OD): Enter the percentage of nonlinear gaussian noise. This noise value will affect optical densities at higher values more than at lower concentrations and reflects the instrumental noise seen at higher optical densities in spectrophotometers. Values of 0.01 - 0.02 are common for the XL-A analytical ultracentrifuge.

Radial Increment: This value cannot be adjusted by the user and reflects the radial stepsize resulting from the column height and the number of datapoints selected for each scan.
Simulated Wavelength: This value has no effect on the calculations, but is used when the data is exported to an external format.
Export as: XL-A/UltraScan: Select the export format for the simulated data. "XL-A" format produces ASCII files of the type generated by the Beckman data acquisition software (00001.ra1, 00002.ra1, ...), while "UltraScan" format produces a binary output file suitable for direct analysis with the UltraScan equilibrium modules.

Help: Show this Help file.
Plot: Plot the simulated equilibrium scans in a plot window.
Histogram: Plot a concentration histogram for the total concentration of all scans in the simulated experiment.
Export: Export the simulated data to an external file format. See "Export as:" for additional detail.
Close: Close this module and exit.

www contact: Borries Demeler

The latest version of this document can always be found at:

http://www.ultrascan.uthscsa.edu

Last modified on January 12, 2003.

	Model Selection: To select a model, click on the "Select Model" button. You can then select a predefined or user-defined model from a model selection dialog. Model selection is explained in more detail here.
	Component Information: Species: For multiple component models you can use the arrow button to select the desired component. Mol. Weight (dalton): Enter the molecular weight for the conponent selected above. Concentration (mM): Enter the partial concentration in millimolar for the component selected above. V-bar (ccm/g, 20^oC): Enter the partial specific volume for the component selected above. Clicking on the button will load the Vbar Calculation module, which allows you to estimate a v-bar value from the amino acid composition specified in a protein sequence file. *Ext. Coeff. (OD/cmM):** Enter the extinction coefficient of the component selected above for the the wavelength to be simulated. The units for the extinction coefficient are measured in optical density units at the selected wavelength per cm and mol. Clicking on the button will load the peptide properties calculation module, which allows you to estimate an extinction coefficient for the denatured protein at 280 nm from the amino acid composition specified in a protein sequence file.
	Model Parameters: Assoc. Const. 1/2/3: Enter the natural log of the desired association constant in molar concentration units. Fields will be disabled for models that do not require association constants. Baseline OD: Enter the baseline concentration, if any for the simulated experiment. Loading OD Contrib.: The figure listed in this field is the contribution to the optical density for the selected component. This number is calculated from the molar concentration and the extinction coefficient. This number cannot be changed by the user. If you want to change the value listed here, you need to adjust the molar concentration or the extinction coefficient.
	Conc. Increment (OD): Enter the concentration increment for multiple loading concentrations. For a global equilibrium experiment, at least three loading concentrations should be measured (for example, 0.3, 0.5, and 0.7 OD). The increment should be listed in OD units. # of Conc. Steps: Enter the number of desired concentration steps for this experiment. For example, if three different loading concentrations are desired click on the arrow buttons to select "3". The double arrow buttons chnage the setting in steps of 10, the single arrow button in unity steps. Temperature (^oC): Enter the temperature in degrees Celsius for the experiment. If different from 20^oC, the program will automatically adjust the vbar and density settings to the new temperature. Density (g/ccm 20^oC): Enter the density of the buffer for the simulated experiment. The density is entered in cgs units for the buffer at 20^oC. Clicking on the "Density" button loads the buffer calculation module. Column Height (mm): Select the column height in millimeter for the experiment. Meniscus: Displays the current meniscus position. The meniscus position is automatically calculated from the column height and the cell bottom position.
	Pathlength (cm): Enter the centerpiece pathlength (in centimeters). Speed Start: For a global equilibrium experiment it is recommended to use multiple speeds. The correct speed depends on the molecular weight of the component(s) to be simulated, and are best expressed in values of sigma (reduced molecular weight) ranging between 0.5 and 4. The Equilibrium Speed Estimation module can assist you in determining the best speeds for your experiment. Use this field to enter the lowest speed of the experiment. Use the arrow keys to select the speed. The single arrow key changes the speed in steps of 100 rpm, double arrow keys change the speed in steps of 1000 rpm, and triple arrow keys change the speed in units of 10,000 rpm. Speed Stop: Select the highest speed for this simulation. Speed Steps: Select the number of different speeds used in this simulation. The program will calculate equal speed intervals between the Speed Start value and the Speed Stop value. If only 2 speeds are selected, the Start Speed and the Stop Speed will be used. If one speed is selected, the Start Speed only will be used. Cell Bottom (cm): Enter the cell bottom position in centimeter to define the geometry of the cell. The single arrow button changes the position in units of 1x10<^-3 cm, the double arrow button changes the position in units of 1x10<^-2 cm, and the triple button changes the position in steps of 0.1 centimeter. Maximum OD: Enter the maximum OD to be simulated in the experiment. All data exceeding this value will not be displayed.
	# of Datapoints: Select the number of datapoints for this simulation. Each scan will be simulated with that many points, which will be equally spaced wit the radial step size listed under "Radial Increment". Gaussian Noise (% OD): Enter the percentage of the total optical density to be simulated as random noise, with gaussian distribution. Single arrow button clicks will adjust the noise level in steps of 0.01 OD percentage points, double arrow button clicks will adjust the level in steps of 0.1 OD percentage points, and single arrow button click will adjust it in steps of unity. 0.1 - 0.3 percent of 1 OD are common values for the XL-A analytical ultracentrifuge. Nonlinear Noise (% OD): Enter the percentage of nonlinear gaussian noise. This noise value will affect optical densities at higher values more than at lower concentrations and reflects the instrumental noise seen at higher optical densities in spectrophotometers. Values of 0.01 - 0.02 are common for the XL-A analytical ultracentrifuge.
	Radial Increment: This value cannot be adjusted by the user and reflects the radial stepsize resulting from the column height and the number of datapoints selected for each scan. Simulated Wavelength: This value has no effect on the calculations, but is used when the data is exported to an external format. Export as: XL-A/UltraScan: Select the export format for the simulated data. "XL-A" format produces ASCII files of the type generated by the Beckman data acquisition software (00001.ra1, 00002.ra1, ...), while "UltraScan" format produces a binary output file suitable for direct analysis with the *UltraScan* equilibrium modules.
	Help: Show this Help file. Plot: Plot the simulated equilibrium scans in a plot window. Histogram: Plot a concentration histogram for the total concentration of all scans in the simulated experiment. Export: Export the simulated data to an external file format. See "Export as:" for additional detail. Close: Close this module and exit.