Detecting bands by ImageJ on mac not fixed properly in each lane?

I am using ImageJ (free version on the website)

on mac and when I select bands, the first one detected correctly, when I click cmd 2 (= ctr 2 on windows system) to detect the second band, the rectangle gets fixed on the previous one (which is band number 1) and the same for the rest (attached photo to deliver the question more clear) … I tried to read how to solve this and unfortunately, I couldn’t find an answer… Can any one let me know the reason and how to solve this issue???

Thanks in advance

Hi @rashaaref,
as a welcome gift you will receive a link to the important ImageJ UserGuide

In section 30.13 you will find some instructions how to use the gel analysis tools (or online here).

Most important: Select a lane not a band.

Hope this helps as a first hint.

Thanks for the reply, I am trying to follow this guideline but I can’t get clearly should I do the following step :
“Under the Analyze menu, choose calibrate and open the calibration.txt file (note
black=0 and white =256). This file will convert 0 to black and 256 to white. If
this file is not on your computer then create it:. Next, choose function “straight line” in the pull down menu of the calibration pop
up window. (You may have to enlarge the window by pulling on the lower right
corner to see the OK button). Click OK”
Or leave the default setting??
Thnaks in advance

Where have you found this?
Which guidelines are you using?

Have you done a test with ImageJ sample image gel.gif ?(https://imagej.nih.gov/ij/images/)
What result do you get when following the instructions of the user guide with the gel.gif image?

I have highlighted the parts in question, kindly clarify these issues !!!

DensitometryofWesterenblotsusingIMAGEJ.pdf (7.0 MB)

When I clicked on the provided link I got many links, which link should I use, and what is the purpose behind this link ??? Thanks in advance

I am not sure how my question can be more specific. SORRY !!!