Start ImageJ and open the MISA++ plugin by navigating to Plugins → MISA++ for ImageJ. This will open a list of all available MISA++ applications.

Select “ MISA++ Kidney Glomeruli Segmentation”.

        </p>
        <p >Click
        <img src="/img/imagej/userguide_html_63631d3be680189.png" class="inline-image" name="image80.png" border="0"/>

Launch to open a tool that allows preparation of the data and changing parameters.

        </p>
    </td>
</tr>
<tr valign="top">
    <td ><p>
        <img src="/img/imagej/userguide_html_d51c71947cefea0d.png" name="image128.png" border="0"/>

Click Import folder and select the folder that contains the example data set to automatically add samples and import images.

        </p>
    </td>
</tr>
<tr valign="top">
    <td ><p>
        <img src="/img/imagej/userguide_html_ffb1140b27a42629.png" name="image16.png" border="0"/>

Select the folder that contains our example data set and click Open.

        </p>
        <p>Then delete the default sample “<img class="inline-image" src="/img/imagej/userguide_html_e507c96bf16d7da5.png" name="image141.png" border="0"/>

New Sample”, as we already imported all necessary data.

To enable multi-threading, select the Runtime tab and increase the number of threads.

        </p>
        <p >Click
        <img src="/img/imagej/userguide_html_63631d3be680189.png" class="inline-image" name="image76.png" border="0"/>

Run and then Run now to start the analysis. After the analysis finished, the plugin will ask you to further analyze the results.

        </p>
    </td>
</tr>

The analysis tool displays a list of all output data and (if available) input data and allows importing data back into ImageJ.

If you want to import the glomeruli into ImageJ, click Bioformats Import next to the “glomeruli3d” data.

To analyze quantification results, click Browse quantification results.

        </p>
        <p >This will open a tool to browse
        and filter quantification results and transform them into tables
        that can be plotted.</p>
    </td>
</tr>
<tr valign="top">
    <td ><p >
        <img src="/img/imagej/userguide_html_7c7db2265df6ffa2.png" name="image145.png" border="0"/>

To analyze the glomeruli, select Objects in the middle column and then “ Glomerulus” above the table.

Due to performance reasons, the table does not contain all available information about each glomerulus. To add more, click Edit columns.

        </p>
        <p >Click
        <img src="/img/imagej/userguide_html_96edca7cbbbe705d.png" class="inline-image" name="image118.png" border="0"/>

Clear selection and select the columns

• Sample

• diameter/Value

• diameter/Unit/unit

• Valid

If you are happy with the table, click Analyze.

        </p>
        <p >This will open a tool that allows
        you to modify the table and perform statistics.</p>
    </td>
</tr>
<tr valign="top">
    <td ><p >
        <img src="/img/imagej/userguide_html_6c577a41ad99985a.png" name="image140.png" border="0"/>

To calculate mean and variance of the diameter,

select a cell in the “diameter/value” column and navigate to Add column → Copy selected column. Then click “OK”.

        </p>
        <p ><br/>

        </p>
        <p ><br/>

        </p>
    </td>
</tr>
<tr valign="top">
    <td ><p >
        <img src="/img/imagej/userguide_html_45fc33e19f6e192d.png" name="image136.png" border="0"/>

Click “Integrate columns” at the top right corner and assign following roles to each column:

• Sample: Category

• diameter/unit/unit: Ignore

• diameter/value: Average

• valid: Category

• diameter/value: Variance

Click “Calculate”.

The table now shows the mean and variance of the glomeruli, categorized by the sample and if the glomerulus is valid according to the segmentation algorithm.

        </p>
        <p >Click
        <img src="/img/imagej/userguide_html_a590cddcece250fa.png" class="inline-image" name="image24.png" border="0"/>

Undo to restore the original table and then Create plot.

Set the plot type to Box Plot, set the category to “Table.Sample”, X axis to “Table.valid” and the list of values to the diameter.