Bouin's solution (invented by the French biologist Pol Bouin) is a fixative element composed by saturated picric acid (75%), formol (20%) and glacial acetic acid (5%). Also, be careful to work in a well ventilated area, wear gloves, lab coat and goggles because his use has waned due to safety concerns (NIOSH, 2014; http://www.cdc.gov/niosh/idlh/88891.html).

Small biopsies could be fixed in 2 to 4 hours and large specimens may remain in Bouin's solution up to 3 days maximum. In  general,  low  temperatures  retard autolysis,  but  they  also  decrease  the  diffusion  rate  and  thus  prolong  penetration… For (adult ?) rat testes, a little bit more than an overnight session would be great. After the fixation, you could put the tissue in alcohol 70 at 4°C before any further utilisation.

The use of Bouin's solution depends on the techniques you need to proceed on rat testes. It is especially good for nuclear staining observations (like H&E / HES or PAS). An exemple of seminiferous cord observed on immature rat testicular tissue (using HES staining at ×500 magnification) could be observed in an article published by the group around Nathalie Rives (Travers A. et al., 2011).

However, it was an awful fixative if you need to observe tissue structures on (i) electron microscopy or (ii) fluorescence on IHC (4% formaldehyde at pH = 7.4 is the prefect candidate for this kind of job).

Hoping that this reply will be useful,

Ludovic

http://www.cdc.gov/niosh/idlh/88891.html

Rinsing tissue in 2% lithiumcarbonat until yellow colour is gone. Then continue with 70% ethanol, 80%, 96% 2x, 100%2x, Xylen 2x, paraffin 3x. Time depends on the size of specimen.

If sections are still yellow rinse in tapwater until colourless after deparaffination and rehydration.

Bouins is traditionally recommended for testes, because counting mitosis should be easier. Otherwise there is no real cause, why not fix with 4% NBF, which is more suitable for IHC.

If I use NBF, is it I still can see the mitosis clearly? Before this I use formalin, but I can't clearly differentiate the stage of spermatogenesis. So, I plan to use Bouin but some said that Bouin can cause shrinkage of the cell and can't keep the tissue in Bouin for a long period of time.

The specimens should be transferred to 70% ethanol after fixation for long-time storage. If you have the possibility, fix one half of the testes in Bouin and one half in NBF. So you can make your own decision, what is better for your purpose. 

In pharma research (toxicologic pathology), the fixative of choice for testis is a modified Davidson's solution, which provides better morphology than NBF, without the problems associated with Bouin's. See ref: Toxicol Pathol. 2002 Jul-Aug;30(4):524-33.

After fixation in bouins  fluid thaw the tissue in tap water &transfer it sodium thiosulphate of graded % till the yellow colour is lost. Time taken for each solution of  sodium thiosulphate depends on thicikness of the tissue

After fixation of tissue block in Bouin's fluid for 24 hours, it should be kept in running tape water at least three hours before dehydration process.