Feulgen and Rossenbeck staining

Fig. 1. Quail cells are differentially identified in 5- m paraffin sections of a chimeric cerebellum (see refs. 16,21,22). (A) With the Feulgen and Rossenbeck staining (F R), quail neuronal and glial cells show a strongly stained nucleolus (see arrowheads), whereas chick cells present almost homogeneously stained nuclei. (B) With the QCPN MAb, nucleolus of quail neurons and glial cells are more violently stained, but chick nuclei are not labeled. (C) The SMP probe hybridizes specifically with the quail oligodendrocytes. Medial and high magnifications of each staining are presented. Al, Bl, Cl bar = 7O 0.m A2, B2, C2 bar = 3O 0,m.

Host embryos can be fixed from several hours after the operation to several days after hatching according to the experimental design (see Note 20). Carnoy fluid is one of the best fixatives utilizable at once for Feulgen and Rossenbeck staining (13), immunohistochemistry, and in situ hybridization on paraffin sections. Fixation with 4% paraformadehyde is used for whole-mount immunohistochemistry and in situ hybridization, and all treatments on cryostat sections. 

Zenker fluids like Camoy fluid are good fixatives for Feulgen and Rossenbeck staining (13) but they do not allow immunohistochemistry or in situ hybridization to be made. 

After the desired period of incubation, fix the embryo by flooding with methanol (for most immunological detection procedures), Zenker s fixative (for Feulgen-Rossenbeck staining), or in 4% formaldehyde in PBS (for in-situ hybridization and most other procedures). In the case of methanol or Zenker s, which are rapid fixatives, it is advantageous to submerge the cultured embryo in saline, then detach it from the vitelline membrane and transfer it to a clean dish with saline prior to fixation. Otherwise, the embryo adheres permanently to the membrane and the fixative denatures the albumen, generating threads of protein that tend to stick to the embryo. Formaldehyde can be poured directly onto the embryo, provided that the embryo is detached from the membrane within a few minutes. 

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