Converting and Analyzing ND2 files


I am trying to open images in image J that were saved as ND2 files taken on a Nikon Eclipse Ti. The files are opening up in image j as stacks of images on top of each other. I was wondering if there was a way to either convert all of the ND2 files to .tif without going back to the scope and re-saving as different files, or if there was a way to fix the files in image j so they are back to one channel with RGB. Just as a note, these images were not taken as stacks. They were taken as 2D images.

Thank you,


Dear @LDeniau,

welcome to the forum! :slight_smile:

You can convert your images via Process > Batch > Convert (check the “Use Bio-Formats” box) to TIFF. You can do a conversion from multi-channel to RGB by pasting the macro below into the textbox of Process > Batch > Macro:

run("RGB Color");



Hi @LDeniau,

Do you mean the images open as z stacks rather than composite (multichannel)?

If you want to preserve the channels as separate (essential for quantitation), you can do Image > Hyperstacks > Re-order Hyperstack and choose Channels >> Slices and Slices >> Channels as below. Then your “z stack” will become a multi-channel image.

You can fix the colors using Image > Color > Channels Tool.

I would not use the “RGB Color” conversion if I was going to analyze the intensity or particles later.

Hope this helps.


Ugh, I might have completely misunderstood the question :worried:

Could you clarify the structure of your data in the ND2 files, @LDeniau?