Hi all,
I am doing Rapid Golgi staining for mice cerebellum using the FD NeuroTechnologies kit. I need to to do the dendrite spine qualifications of Purkinje cells . Does anyone know how to use ImageJ for these quantification?
Hi,
first of all, the method of image capturing is important for visualization of spines, by which microscope and method did you capture the image?
Hi Maryam,
I am using 80um and 100 um thick brain slices and 100 X images were taken from conforcal microscope
Great! what is your plan to analyse spines? whole neuron or selected branches? I am asking due to the complex morphology of Purkinje cells , I recommend you IMARIS software, flament tracer module.
Hi Maryam,
Thanks for the recommendation. I am planing to analyse morphological changes in spines and spine density under different conditions. It seems that t will be much difficult to quantify an entire Purkinje cells. So my plan is to do the quantification on some selected branches in periphery.
Hi Lahiru,
Imaris is a great option, but if you don't have a full licence, you can use free tools such as Vaa3d, neuTube and TREES Toolbox. These tools allow you trace the morphology and create tree structures on which you can run many types of analysis. You can also separately identify the spines using these tools.
Hi Sumit,
Thanks for the suggestion. It is really useful. I will try both Imaris trial version and the tools you suggested.
Hi Lahiru,
May I ask which analytical method you ended up going with? Im also currently trying to find the most efficient way to analyze dendritic spines using golgi stain.
Hi Nicholas Deems ,
I took the images as described above, but the quantification we did manually, since I could not get the Imaris licence version and other software did not work with me very well.
As mentioned above, we decided to select some branches in periphery of Purkinje cells and went with branching points, ration between mature and immature dendrite spins and spins density.
Good luck
It seems quite difficult to make 3D reconstruction for Golgi-stained samples. Because the stained dendrites can absorb transmission light, so the spines behind dendrite are shadowed, and the shape of dendrites are not in cylinder shape.
It's very difficult to quantify by using Rapid Golgi Method because of the silver impregnation isn't complete. I recommend you the Cox Golgi method for complete impregnation and for cuantification the software Reconstructor from Texas University is a nice help
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