how to merge lsm file withe a picture

[Fabiano Perina]

Hi everyone,

I've been tried put two images differents of the same sample, where one is from
a Nikon D7000 digital camera (.jpg file) take with a macro lens 60 mm and another image is a .lsm file form a High speed confocal microscope and contains a z stack in two color (red and cyan, where I have 14 slices per channel).

The problem that I have to face is

1) First how the better way to do it? (I've tried use first split channels, follow by merge channels, make a 3d projection....)

2)The big problem is: even if I change the image from digital camera to the same pattern  (width, hight, voxel depth,  canvas size, etc.) I still can not put the images together because the useful size (from digital camera image) keeps small than the microscope confocal images (lsm files that contains 17 slices in each channels).

How can do this do to merge and to create a animated in order to have a Idea about the picture and the micrographs from confocal in the same video or animation file? Please anyone have some ideas about it?

I would be very greatful if you could help me and I would greatly appreciate your help!

With my best regards, Perina

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[Pariksheet Nanda]

On Sun, Nov 4, 2012 at 4:48 PM, Fabiano Perina <[hidden email]> wrote:
> I've been tried put two images differents of the same sample, where one is from
> a Nikon D7000 digital camera (.jpg file) take with a macro lens 60 mm and another image is a .lsm file form a High speed confocal microscope and contains a z stack in two color (red and cyan, where I have 14 slices per channel).
>

It seems like you're trying to overlay your confocal fluorescence
images with (presumably) transmitted light from your D7000.
it would be easiest if you could setup your confocal microscope
software to acquire that transmitted light image while collecting the
confocal dataset.
Your software might have control over the transmitted light shutter or
be able to toggle the lamp voltage or on/off state.

> With my best regards, Perina

Pariksheet

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[Christian Goosmann]

> Hi everyone,
>
> I've been tried put two images differents of the same sample, where one is from
> a Nikon D7000 digital camera (.jpg file) take with a macro lens 60 mm and another image is a .lsm file form a High speed confocal microscope and contains a z stack in two color (red and cyan, where I have 14 slices per channel).
>
> The problem that I have to face is
>
> 1) First how the better way to do it? (I've tried use first split channels, follow by merge channels, make a 3d projection....)
>
> 2)The big problem is: even if I change the image from digital camera to the same pattern  (width, hight, voxel depth,  canvas size, etc.) I still can not put the images together because the useful size (from digital camera image) keeps small than the microscope confocal images (lsm files that contains 17 slices in each channels).
>
> How can do this do to merge and to create a animated in order to have a Idea about the picture and the micrographs from confocal in the same video or animation file? Please anyone have some ideas about it?
>
> I would be very greatful if you could help me and I would greatly appreciate your help!
>
> With my best regards, Perina
>
> --
> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>

First you could make a z projection of your confocal data so that you will only have to combine two 2D images. Before combining you could manually scale (and rotate) the smaller image so that it matches the scale of the larger one. For combining you could use Image>Overlay>Add Image... command in ImageJ making the overlay semitransparent.
If you want it in 3D, you could change the Nikon image in size so that it matches the scale of the confocal data, rotate and crop to the x/y size of the confocal stack and add the image to the stack as an additional slice. (use Image>Stack>Add Slice to add an empty slice and copy/paste the image there) Of course the image and the stack need to be the same image type first, so merge the cofocal stack to RGB if the Nikon image is RGB. After doing that you can visualize the combined stack with one of the 3d viewing tools like Image>Stacks>3D Project... etc.
If you have the time and energy to go through some tutorials you should maybe look at the TrakEM2 plugin that lets you load a stack, add an additional layer, rotate, scale etc. that layer for registration with the rest of the stack and finally send the whole thing to 3D-viewer.
I'm sure that these are not nearly all methods you could use.
Good luck
Christian

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