Tips and tricks for using ImageJ

Question

[LauferVA]

Hey Biostars,

Using ImageJ for the first time, in this case to quantify Western Blot data. For reproducibility, I will be using the web-based version for this post. However, if there are relevant features in the standalone versions, please let me know as I would be willing to switch versions.

My basic procedures follows NIH recommendations for use of ImageJ. Briefly,

1. Upload the image
2. Select Edit, then `Invert` the image (such that white --> black; black --> white)
3. Select Rectangle Tool
4. Draw a first rectangle around the first band in lane 1. 
5. Go to Analysis --> Gels --> Select first lane.
6. Move rectangle to second band --> Select next lane.
7. to N. Repeat until every band in the first lane is selected.
N + 1. Click on the first rectangle (from Step 5.). Press the `m` key to measure.
(N + 2). to [(2N + 3) - 7]. Select each rectangle, and press `m` each time.
[(2N + 4) - 7]. Finally, export all the measurements to a .csv file
Finally, start over at step 4. for lane 2 and repeat above steps starting from 4.

Automating this entire process would be best, but strikes me as a bit more involved than I have time for at present for a number of reasons. However, there are a few key steps that if those could be streamlined, the process would become very rapid. My immediate goal for this post is really to speed these few steps up (below), unless there is an easy generalized solution..

Here, I've just completed the process and exported the results to .csv (I am about to restart at step 4 for lane 2).

is there an easy way to move all the rectangles downward the same number of pixels at the same time? In many programs, something like Shift + Click or Command + Click would allow one to select multiple rectangles, for instance ...

Second, when doing the measuring step, currently I must click on each rectangle individually, then hit m, then manually select the next one, press m again, etc., like so:

Is there a way to perform the measurement step for all the rectangles in the lane at once?

Thanks very much for reading. I'd be happy to have answers in the form of hotkeys, macros, or entirely different solutions. VAL

Answer 1

[Jeremy]

Using the web version of ImageJ in Linux, you can use Shift + Click to make multiple rectangles. You can then click and drag those rectangles to move them around as a group. To measure multiple rectangles at once, use the following:

Analyze > Tools > ROI Manager > More > Split

Then:

More > Multi Measure > OK

I haven't used ImageJ in about seven years, so I would use some sample data to make sure that the results of this workflow make sense.

[LauferVA]

was going to post this actually - same series of steps works on the browser version.

did you stop using imageJ beacuse there is a better program, or do you just not need to solve a problem like this anymore?

VAL

[Jeremy]

I'm just not quantifying western blots anymore. I'm not aware of a better program, but there could certainly be one out there.