Testis visualization

CNG channels are expressed in retinal photoreceptors and olfactory neurons, and play a key role in visual and olfactory signal transduction. In addition, CNG channels are found at low density in some other cell types and tissues such as brain, testis, and kidney. While the function of CNG channels in sensory neurons has been unequivocally demonstrated, the role of these channels in other cell types, where expression has been observed, remains to be established. Based on their phylogenetic relationship, the six CNG channels... 

Testis squashes can be stained with antibodies directed to specific proteins, with rho-damine-phalloidin to visualize F actin, and with DNA-binding dyes. Best results are obtained when these stainings are carried out in the following order immunostaining, F-actin staining, and DNA staining. This implies that multiply stained preparations are all mounted in the media described for DNA staining (see Protocol 5.8, step 2). 

Several techniques have been developed for preparation of male meiotic chromosomes (see, e.g.. Cooper 1965 Ashbumer 1989 Miyazaki and Orr-Weaver 1992). All of these techniques allow a clear visualization of chromosomes, but not of other cellular structures such as the cytoskeleton and the spindle. We describe here three of these techniques that in our hands give good reproducible results. The first technique, developed by Lifschytz and Hareven (1977) and described below in Protocol 5.9, is particularly usefid for the analysis of adult testes. The other two techniques (Ripoll et al. 1985 Cenci et al. 1997) are routinely used in our laboratory for larval and pupal testis preparations. The technique described in Protocol 5.10 is a very simple method for preparing aceto-orcein-stained chromosomes. The other technique outlined in Protocol 5.11 allows preparation of unstained chromosomes that can be subsequently stained with various dyes such as Giemsa, Hoechst 33258, and DAPl, or processed for in situ hybridization. 

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