The operation manual of spotlight 300 IR microscope Make sure there is no sample under the microscope and then click spotlight on the desktop to open the software. You can do imaging with the image mode and line scanning and mapping with the point mode. Imaging NOTE: Spotlight must be in Image mode before you can scan an image. To switch to Image mode select Image in the Control window. 1. Load the sample. In the Stage Control window, click, or select Load Position from GoTo menu in the Stage Control Window, to drive the sample stage to the loading position. Position the sample in the center of the sample stage. Then click, or display the GoTo menu and select Center of Stage to drive the sample stage so that the sample is in the center. 2. Look at the sample and focus it. The Monitor Visible window displays a visible image. To focus this image, use the controls on the Imaging System. 3. Collect a visible image survey. The Monitor Visible window displays only the field of view of the Imaging System, which is a small area. Select a default survey size from the View menu in the Stage Control window. (To get an overall picture of your sample, click in the Stage Control window.) A visible image is collected and displayed in the Stage Control window. 4. Collect an image background. Using the joystick, move to a clear area that you wish to use for the background. In the Stage Control window, click. Check the settings in the Scan Image Background dialog and click Start to begin scanning. 5. Scan the image. Using the joystick, move back to the region you want to scan. In either the Monitor window or the Stage Control window, draw a grow box around the area you want to scan. In the Stage Control window, click. The Start Image dialog is displayed. Enter a filename for the new image and click Start Image. Data collection begins and the new image is displayed in a new window. Point Mode In Point mode, you can collect individual spectra (either at the current position or at positions you have marked). You can also collect spectra at regularly spaced intervals along a line (line-scanning) or within a marked area (mapping). In Point mode, you can choose whether the spectra you collect are in ratio or single-beam mode. If you choose ratio mode, the sample spectra are automatically ratioed against a background spectrum, so you must collect a background spectrum before you begin scanning (see Collecting a
Background Spectrum in Point mode). However, if you collect spectra in single-beam mode, you can ratio them against a background spectrum afterwards, (see Ratioing Single-beam Spectra). on this marker In Point mode, you can change the size and shape of the infrared aperture. The red box which is displayed at the current position represents the current aperture (see Setting the Infrared Aperture). Double-clicking displays the Infrared Aperture dialog. Collecting a Background Spectrum in Point Mode If you want to collect spectra in ratio mode, you must first collect a background spectrum. 1. Move the sample stage so that the area on the sample from which you want to collect spectra is in the current aperture. 2. Set the infrared aperture. For mapping, it is usual to set the aperture to the size of the smallest feature (see Setting an Aperture). Setting an aperture does not apply for Imaging. If you are using Spotlight in Image mode, refer to Collecting an Image Background. 3. Move the current position to the area of the sample that you want to use to collect the background spectrum. In the Stage Control window, display the Options menu and choose Set Background Position. This position is marked as the background position. 4. If you are working in transmittance you must maximize the energy reaching the detector, see maximizing the energy reaching the detector in transmittance mode. If you are working in reflectance, you do not need to maximize the energy provided that the infrared focus is the same as the visible focus. 5. In the Stage Control window, display the Scan menu and choose Background or click on. The Scan Background dialog is displayed. 6. Choose a Display Mode.
If you choose Overlay, the spectrum is displayed in an existing view window, if you choose New Window, the spectrum is displayed in a new view window 7. Type the Scan Parameters that you require (see Scan Parameters). 8. If the ATR is required, select Use ATR. 9. Choose OK. If the sample stage is not already at the background position, a dialog is displayed. 10. Choose Yes to collect the background spectrum at the position you previously set, or No to collect the background spectrum at the current position. If the ATR is in use, dialogs will prompt you to raise or lower the ATR as required. When the FT-IR has finished scanning if the sample stage has moved to collect the background spectrum, it returns to its previous position. The Imaging System returns to view mode and the resulting background spectrum is displayed. Again, if the ATR is in use, dialogs will prompt you to raise or lower the ATR as required. Collecting a Spectrum at the Current Aperture Position 1. Move the current position to the part of the sample where you want to collect a spectrum. 2. In the Stage Control window, display the Scan menu and choose Sample Ratio or Sample Single Beam, depending on whether you want to collect spectra in Ratio Mode or Single-beam Mode. OR Click on (ratio) or (single-beam). The Scan Sample Ratio or Scan Sample Single dialog is displayed. The Imaging System automatically changes to IR mode, and the Monitor Visible window becomes black. 3. Choose the Display Mode. If you choose Overlay, the spectrum is displayed in an existing view window, if you choose New Window, the spectrum is displayed in a new view window. 4. Type the Scan Parameters that you require. 5. If you are working in transmittance you must maximize the energy reaching the detector, see Maximizing the Energy in Transmittance.
6. Choose OK. When the FT-IR has finished scanning, the results are displayed and the Imaging System returns to view mode. Defining a Line Scan When you have decided on your scan parameters, and on the position of the line, you are ready to define the line scan. 1. In the Stage Control window, use the mouse to zoom in on the area where you want to position the line (see Rescaling the Stage Control Window). 2. In the Stage Control window, display the Edit menu and choose Setup Line Scan. Note: The Setup Line Scan command is not available when Spotlight is in Image mode. Switch to Point mode before Line Scanning. The Setup Line Scan dialog is displayed. 3. Choose Define LineScan. The Define Line Scan dialog is displayed. While the Define Line Scan dialog is displayed, you can use the mouse to change the size and orientation of the line. 4. Choose New. A new line scan marker is displayed, which crosses the central part of the Stage Control window, and which has X and Y intervals that are the same as the current X and Y dimensions of the infrared aperture. 5. Use the mouse to position the line scan marker on the part of the sample that you want to scan, and to change its size so that it crosses the features that you are interested in. You can move and size the line scan marker in exactly the same way as a Section Locator. Use the Monitor Visible window to position the line scan marker - its relation to visible features can then be seen clearly. The coordinates of the line scan and the number of points are updated in the Define Line Scan dialog. Notice that the interval between scan points stays the same when you change the size of the line.
6. Type values for either the Interval between scanning points or the Number of Points in each direction, and choose Update in the appropriate box. The line is updated to show the new intervals between line scanning points. The Update button that you choose affects the spacing of the scan points. If you choose Update in the Interval section, the interval between points stays the same, and the number of points on each axis is recalculated. If you choose Update in the Number of Points section, the number of points on each axis stays the same, and the interval is recalculated. 7. When the line is defined as you require, press OK. The Setup Line Scan dialog is displayed. 8. Choose Ratio or Single Beam mode. 9. Type the Scan Parameters that you require (see Scan Parameters). 10. If you want to automatically collect new background spectra at regular intervals during line scanning, choose Scan background every, and type a number of points in the text box. 11. If you want to automatically refocus the sample at regular intervals during line scanning choose Autofocus every, and type a number of points in the text box. If you choose Autofocus every, at the interval you have set: the Imaging System changes to visible mode; the image in the Monitor Visible window is automatically focused; the Imaging System changes to infrared mode; line scanning continues. 12. If you are collecting an ATR line scan using the ATR accessory, choose ATR Mode. See ATR Line Scanning 13. Choose OK. You are now ready to start the line scan. Starting a Line Scan
1. In the Stage Control window, display the Scan menu and choose Start Line Scan. Note: The Start Line Scan command is not available when Spotlight is in Image mode. Switch to Point mode before Line Scanning. The Start Line Scan dialog is displayed. The Imaging System automatically changes to IR mode, and the Monitor Visible window becomes black. 2. Type a filename for the results, or choose Select and choose a file for the results. 3. If you want to monitor the energy before you start line scanning, choose Monitor; if you want to scan a new background, choose Background. 4. Choose Summary. 5. The Line Scan Summary dialog is displayed. Make sure that the line scan setup is correct. This dialog contains a Print button, which enables you to print the summary information. 6. Choose Close. The Line Scan Summary dialog closes. 7. Choose OK in the Start Line Scan dialog. The Line Scan Information dialog is displayed. 8. If the line scan duration is too long or the file size is too large, choose Cancel, and change one or more of the settings that affect scanning time and file size. The scan parameters (Setup Line Scan dialog), the number of points (Define Line Scan dialog) and the FT-IR settings all affect the scanning time (see Scan Parameters and FT-IR settings). The Imaging System automatically changes to IR mode, and the Monitor Visible window becomes black. 9. If you want to be able to see the visible image while you are line scanning, survey the visible image first. Line scanning starts, and the progress is indicated by blue crosses, which appear on the line when each spectrum has been collected. 10. To halt the line scan before it is completed, display the Scan menu and choose Halt.
OR Click on. The data that has been collected so far is saved. The Imaging System returns to view mode. When the line scan is completed a message box is displayed and the Imaging System returns to view mode. 11. Choose OK. A 3D view of the data is displayed. Defining a Map 1. In the Stage Control window, use the mouse to zoom in on the area that you want to map (see Rescaling the Stage Control Window). 2. In the Stage Control window, display the Edit menu and choose Setup Map. Note: The Setup Map command is only available when Spotlight is in Point mode. The Setup Map dialog is displayed. 3. Choose Define Map. The Define Map dialog is displayed. While the Define Map dialog is displayed, you can use the mouse to change the size of the map boundary. 4. Choose New. A new map boundary is displayed, which fills the central part of the Stage Control window, and which has X and Y intervals that are the same as the current X and Y dimensions of the infrared aperture. 5. Use the mouse to position the map boundary on the part of the sample that you want to map, and to change its size so that it includes the features that you are interested in. You can move and size the map boundary in exactly the same way as a grow box. Use the Monitor Visible window to position the edges of the map boundary - the relation of the map boundary to visible features can then be seen clearly.
The coordinates of the map boundary and the number of points are updated in the Define Map dialog. Notice that the interval between scan points stays the same when you change the size of the map. 6. Type values for either the Interval between scanning points or the Number of Points in each direction, and choose Update in the appropriate box. The map is updated to show the new intervals between mapping points. The Update button that you choose affects the spacing of the scan points. If you choose Update in the Interval section, the interval between points stays the same, and the number of points on each axis is recalculated. If you choose Update in the Number of Points section, the number of points on each axis stays the same, and the interval is recalculated. 7. When the map is defined as you require, press OK. The Setup Map dialog is displayed. 8. Choose Ratio or Single Beam mode. For ratio mode, you must have a valid background spectrum. 9. Type the Scan Parameters that you require (see Scan Parameters). 10. If you want to automatically collect new background spectra at regular intervals during mapping, choose Scan background every, and type a number of points in the text box. 11. If you want to automatically refocus the sample at regular intervals during mapping choose Autofocus every, and type a number of points in the text box. If you choose Autofocus every, at the interval you have set: the Imaging System changes to visible mode; the image in the Monitor Visible window is automatically focused; the Imaging System changes to infrared mode; mapping continues. 12. If you are collecting an ATR map scan using the ATR accessory, choose ATR Mode. See ATR Mapping. 13. Choose OK.
You are now ready to start the map. Starting a Map 1. In the Stage Control window, display the Scan menu and choose Start Map. Note: The Start Map command is only available when Spotlight is in Point mode. The Start Map dialog is displayed. The Imaging System automatically changes to IR mode, and the Monitor Visible window becomes black. If you want to be able to see the visible image while you are mapping, transfer the visible image to a new view window (see Visible Images). 2. Type a filename for the results, or choose Select and choose a file for the results. 3. If you want to monitor the energy before you start mapping choose Monitor; or if you want to collect a new background spectrum, choose Background. 4. Choose Summary. The Map Summary dialog is displayed. 5. Make sure that the map setup is correct. This dialog contains a Print button, which enables you to print the summary information. 6. Choose Close. The Map Summary dialog closes. 7. Choose OK in the Start Map dialog. The Mapping Information dialog is displayed. 8. If the map duration is too long or the file size is too large, choose Cancel, and change one or more of the settings that affect scanning time and file size. The scan parameters (Setup Map dialog), the number of points (Define Map dialog) and the FT-IR settings all affect the scanning time (see Scan Parameters and FT-IR settings). Mapping starts, and the progress is indicated by blue crosses, which appear on the map when each spectrum has been collected. 9. To halt the map before it is completed, display the Scan menu and choose Halt.
OR Click on. The data that has been collected so far is saved. The Imaging System returns to view mode. When mapping is completed a message box is displayed. 10. Choose OK. Default views of the data are displayed. The default views are of a Total absorbance image displayed in false color format, with a crosshairs cursor linked to a spectrum view (see View Systems).