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Protein Quantification Using ImageJ: Difference between revisions
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→Determining the concentration of protein in SDS-PAGE gel bands using ImageJ
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* Does your image look too dark or too light? | * Does your image look too dark or too light? | ||
:- '''Image→Adjust→Brightness/contrast''' | :- '''Image→Adjust→Brightness/contrast''' | ||
* If image looks good... | :- I recommend saving the image with an updated name at this point so that you have it to go back to | ||
* If image looks good as is... | |||
4. Find the lane with the lowest concentration of BSA | 4. Find the lane with the lowest concentration of BSA | ||
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6. Go to '''Analyze→Gels→Select first lane''' | 6. Go to '''Analyze→Gels→Select first lane''' | ||
:- Can also use key command "Ctrl+1" | |||
:- A tiny “1” will appear in the lane | :- A tiny “1” will appear in the lane | ||
:[[Image:Imagej1.jpg|200px]] | :[[Image:Imagej1.jpg|200px]] | ||
7. Make sure your cursor shows as an arrow, grab the rectangle you just made, and drag it to the next lane | 7. Make sure your cursor shows as an arrow, grab the rectangle you just made, and drag it to the next lane | ||
:- '''DO NOT DRAW A NEW RECTANGLE! You must drag the same rectangle you just made | :- '''DO NOT DRAW A NEW RECTANGLE!''' You must drag the same rectangle you just made | ||
:- | :- The point here is to compare the band in each subsequent lane using the exact same size/white space/noise as the originally defined area in Lane 1 | ||
8. '''Go to Analyze→Gels→Select next lane''' | 8. '''Go to Analyze→Gels→Select next lane''' | ||
:- Can also use key command "Ctrl+2" | |||
:- A tiny “2” will appear in the lane | :- A tiny “2” will appear in the lane | ||
:- [[Image:imagejlane2.jpg|200px]] | :- [[Image:imagejlane2.jpg|200px]] | ||
9. Repeat steps 7-8 until all lanes have been selected and numbered | |||
:- Use key command "Ctrl+2" to continue numbering the subsequent lanes | |||
:- I do not know how to correct the inevitable mistake boxes that you are going to make that cannot be undone. Deleting them will cause white space where the rectangle was previously. My best advice if you find yourself in that predicament is to close the file, reopen it and start again. You will get really good at marking the lanes, I promise. If you are some kind of wizard who knows how to fix these mistake boxes without having to start over again, you should let me know! | |||
