First, apologies if this question has been answered in the archives - I couldn't find anything that matched exactly what I want to know.
I am trying to find a way to image the branchial arches of a midgestation mouse embryo immunofluorescently stained with up to 3 antibodies + DAPI
counterstain. Branchial arches are structures on the "neck" of the embryo and are about 400um in the shortest dimension
at the time I want to image.
I need to take a 3D look to see the patterns of cells and how they overlap. It needs to be high enough resolution to see colocalisation of antibodies in individual cells. My staining works best at 15-20um.
So, my question: Is the best approach to take 20 sections at 20um (approximately 400um total), stain them, take a z-stack of each section and then align/register them (with e.g. ImageJ/Fiji)?
I have access to a Leica TCS SP2 confocal laser scanning microscope which can perform z-stacks of approx 80um.
Also, generally does anyone have experience of this kind of experiment and, if so, do you have any tips for getting a nice reconstruction at the end?
Finally, if you know of any good guides or papers out there which have attempted similar things they would be extremely helpful.
Thanks in advance!