Fiji - importing .nd2 files and exporting them with preserved virtual focus (planes/stacks)

I am new to Fiji and scientific image editing (helping a scientist post scans of microscopic slide online) and would appreciate if someone can give me step by step instructions regarding following situation:

  • I have .nd2 files that need to be converted in such a way to preserve all characteristics of a stacked image and be very similar ( the same) as what students would see using a microscope. I used Bio-formats plugin to import the image into Fiji, but am not sure what the next step should be. If I just export it as a tiff (using Bio-formats) it flattens the image. How do I ‘unzip’ .nd2 image into stacks and what do i do to export it in a correct way?
    Thank you very much for the help,

Hi Daniela,
The Bio-formats importer shouldn’t flatten the image. I regularly use it to open .nd2 images and have never seen it flatten an image. Depending on the display mode and the image file dimensions (xyc, xyzc, etc) , the image may be displayed as a composite, in which case all channels are combined to create a multi-coloured image. However, the individual channels are still retained and you can switch the focus between the individual channels. You can also use the channel tool (Image->Color->Channels Tool) to change the display mode and turn individual channels on/off. If your images are opened in composite mode, note that there is a slider (or 2 if you also have z slices) at the bottom of the image. Moving the slider doesn’t appear to change the image, but note that the text at the top of the image changes colour indicating which channel has the current focus.
All of this is usually retained when saving the files as tiff files.
In order to be more specific it would be useful if you could provide a sample image and the precise Bio-formats importer options you are using.
Hope this helps,

Thank you Volko - I believe I am doing everything correctly. (Importing .nd2 with Bio-formats and exporting it into .tiff again using Bio-formats). I do have two sliders at the bottom of the image. What is the best way to present these images online and retain virtual focus? From everything you told me, it looks like this is not the problem with Fiji export but with the in house built web space where I am uploading these images…

Sorry, I don’t really have any experience with displaying multi-channel tif files in a web space and retain the ability to move through the slices.
Perhaps you could use ImageJ as a web application. There is an old post on the forum that may be of relevance (Running ImageJ within a web page), but again that is something I have never tried. Perhaps somebody else can help with that.
Good luck,