Difference between revisions of "Optical Microscopy: Part 2 Report Outline"

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(Created page with "* Update the apparatus section of your report to reflect the changes you made in part 2. <br /> <ol start="4"> <li>Fluorescence imaging and flat-field correction <ol> ...")
 
 
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* Update the apparatus section of your report to reflect the changes you made in part 2.
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# Microscope documentation
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<br />
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## Include an updated block diagram of your microscope.
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# Images
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<ol start="4">
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## Include a figure with an images of the 3.26 &mu;m fluorescent microsphere samples, and the stained cell samples with and without Cyto-D.
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  <li>Fluorescence imaging and flat-field correction
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##* For each sample, create 1 figure with 5 panels.
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    <ol>
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##* The panels of the figure should be: A) unprocessed image; B) reference image; C) dark image; D) flat-field corrected image; and E) histogram.  
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      <li>Procedure
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##* In the caption, specify the exposure and gain settings. Each image should have a scale bar. State the dimension of the scale bar in the caption.
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        <ul><li>Document the samples you used and how you captured images (camera settings, software used, etc…)</li></ul>
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##* For panel E, plot histograms of the unprocessed, dark, reference, and corrected image on the same set of axes. Plot <tt>log10( count )</tt> on the vertical axis and intensity on the horizontal axis. Use a line plot instead of a bar chart for the histogram.
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      </li>
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## Image profile
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      <li>Data</li>
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##* For one reference, dark and cell image set, plot an intensity profile across the same diagonal. You may also use a bead image, along with it's unique reference and dark images. The intensity of your three images should be on the same scale, i.e., 0 to 65,535 or 0 to 1. Place all three profiles on a single set of axes for comparison. (Use the <tt>improfile</tt> command in MATLAB.)
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        <ul>
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# Discussion
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          <li>Include raw images of the stained cell sample, reference image, and dark image.</li>
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## How did your beam expander design affect your images?
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          <li>It may be clearer to show the reference image as a surface plot.</li>
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## What differences did you observe between the cells with and without CytoD?
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        </ul>
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      <li>Analysis and Results</li>
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        <ul>
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          <li>Plot intensity profiles of the reference, dark, and raw images on a single set of axes. (Use the <tt>improfile</tt> command in MATLAB.)</li>
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          <li>Use the reference and dark images to correct the raw image for nonuniform illumination and include the corrected image.</li>
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          <li>Include histograms of the sample, reference, dark, and corrected images on a single plot of log(count) vs intensity.</li>
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          <li>Document the steps you used to process the image.</li>
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        </ul>
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      <li>Discussion</li>
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        <ul><li>How did your beam expander design affect your images?</li></ul>
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    </ol>
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  </li>  <li>Resolution
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    <ol>
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      <li>Procedure
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        <ul><li>Document the samples you used and how you captured images (camera settings, software used, etc…)</li></ul>
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      </li>
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      <li>Data</li>
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        <ul>
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          <li>Include an image of the PSF sample with the beads used for resolution measurement indicated.</li>
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        </ul>
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      <li>Analysis and Results</li>
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        <ul>
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          <li>Report the resolution you measured. Make sure to include N and a measure of uncertainty.</li>
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          <li>Show sample Gaussian fits.</li>
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          <li>Include a thorough discussion of error sources.</li>
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        </ul>
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      <li>Discussion</li>
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        <ul>
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          <li>Include a thorough discussion of error sources.</li>
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          <li>Compare the measured value to the theoretical value.</li>
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        </ul>
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    </ol>
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  </li>
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</ol>
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Latest revision as of 18:29, 30 September 2015

  1. Microscope documentation
    1. Include an updated block diagram of your microscope.
  2. Images
    1. Include a figure with an images of the 3.26 μm fluorescent microsphere samples, and the stained cell samples with and without Cyto-D.
      • For each sample, create 1 figure with 5 panels.
      • The panels of the figure should be: A) unprocessed image; B) reference image; C) dark image; D) flat-field corrected image; and E) histogram.
      • In the caption, specify the exposure and gain settings. Each image should have a scale bar. State the dimension of the scale bar in the caption.
      • For panel E, plot histograms of the unprocessed, dark, reference, and corrected image on the same set of axes. Plot log10( count ) on the vertical axis and intensity on the horizontal axis. Use a line plot instead of a bar chart for the histogram.
    2. Image profile
      • For one reference, dark and cell image set, plot an intensity profile across the same diagonal. You may also use a bead image, along with it's unique reference and dark images. The intensity of your three images should be on the same scale, i.e., 0 to 65,535 or 0 to 1. Place all three profiles on a single set of axes for comparison. (Use the improfile command in MATLAB.)
  3. Discussion
    1. How did your beam expander design affect your images?
    2. What differences did you observe between the cells with and without CytoD?
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