Methods of fixing, sectioning and staining amphibian eggs for cytological study.

A combination of methods for fixation (sublimate, cobalt nitrate, formaldehyde, acetic acid in water), inclusion (celloidin dissolved in methyl salicylate, paraffin-paraplast) and staining was used to make serial sections easy, to avoid clefts and to give a good picture of segmentation mitoses, as well as a good contrast of yolk and cytoplasmic components. Four methods of staining were used concerning the Urodele eggs: Safranin-methyl blue-orange G, safranin-picro-blue black naphthol (Curtis), safranin-violet crystal-orange G (Flemming) and Feulgen-methyl blue-orange G. The achromatic apparatus of the normal segmentation mitoses is clearly delineated and the relationships between astral fibers and yolk are different at metaphase and anaphase. By these methods, particularly suitable for demonstration of nuclei, cytoplasm and achromatic apparatus, the cleaving egg may be used as a test for the inhibition of achromatic apparatus and chromosome damage by antimitotic substances. The contrast between vitelline cytoplasm and cytoplasmic non-vitelline abnormal fibrillar systems, produced by transformation of astral and diastematic fibres, is made particularly evident by these methods of staining.