Cell Outlining in Nih-Image

NOTE ON OPENING FILES USING SCION IMAGE

If you have 8-bit images, you can use File…Open

Otherwise you go to File…Import and choose the specifications. The image however cannot be compressed. If image is compressed, save it in Metamorph as a .tiff file with a different name. Metamorph will remove the compression.

CELL OUTLINING IN NIH-IMAGE

1. Open NIH Image/Scion Image icon on the desktop

2. Under Options click on Preferences and make sure Invert Y Coordinates is checked

3. Open up a cell image file

4. Reduce the window size. Use the HAND tool to center the cell in the window

5. Click on the HEART-SHAPED tool for tracing

6. Trace a continuous outline of the cell. Try to get as close as possible to the original point. The tracing can be done either by a mouse or the drawing tablet.

7. Go to File, Export..., and click on XY Coordinates

8. Change the file type to text.

9. Name the file according to convention, adding ".txt" to the end of the file name. Must do this even with step 8 above.

10. Repeat for the cell nucleus

DRAWING OUT CELL OUTLINES USING MATLAB

1. The name of the program is called NIHoutline.m and is in Wonglab7 under C:/Quynh

2. Just follow the directions and you are good to go.

3. CAUTION: If you need to print out the cell outline using Matlab a second time, use the file cell_outline.m instead of NIHoutline.m

****At the time, there is no way to convert to 8-bit images using NIH-image ****

****Also you cannot seem to open up 16-bit images. You must first convert them to 8-bit using Metamorph or rather DO NOT COMPRESS IMAGE during acquistion*****

CELL AREA USING NIH IMAGE

1. Open up the cell image using one of the methods described above

2. Using that Heart Shaped Tool, outline the cell

3. Under Analyze, click on Measure and the area will be displayed on the Info box on the left bottom

4. If you want to save it to a file, then go to File, Export..., and click on Measurements

8. change the file type to text

9. Name file adding ".txt" to the end of the file name

CELL CENTROID USING NIH IMAGE

1. Open the desired files using one of the methods described above.

2. To create a stack, go to Stacks…Windows to Stacks

3. To adjust the position of the picture, use the Hand Tool in the Tools Menu

4. Bring up the results window by going to Analyze…Show Results or press Ctrl+2

5. Go to Analyze…Options and click on X-Y Center

6. Choose the CrossHair Tool and click on the centroid of the cell and the X-Y coordinates should appear in the results window

7. Continue with the rest of the pictures in the stack

8. Once data has been logged, go to File…Export and a menu will appear. In this menu, make sure Measurement is checked, File Type is text and once again, when naming the file, you have to add “.txt”

9. The file is saved as a text file and you can copy and paste to Excel if need be

Contact Quynh with any questions.