340 VITAL STAINING 



762. Staining with Janus Green in vitro. Tissue Cultures. With 

 cover-glass cultures, raise the coverslip, and fill the well of the 

 hollow slide with the dye solution. Strangways (Cambridge, 1924) 

 recommended 1 in 40,000 dilutions. The culture can be transferred 

 to the incubator for five to ten minutes, but is best examined 

 under the microscope on a warm stage, or in a box maintained at 

 body temperature. 



763. Fixation of Vitally Stained Mitochondria. No method has 

 yet been devised which is generally aj^plicable to cells vitally 

 stained with Janus green, or the other Janus dyes. Temporary 

 fixation of the dye is possible in isolated cells, or in cells growing 

 in tissue cultures, by Lewis method {Amer. J. Anat., xxx, 1922, 

 p. 39). With tissue cultures Lewis advises raising the coverslip 

 and placing a small flake of iodine at the bottom of the well of the 

 hollow slide. With isolated cells teased out on a slide, A few 

 flakes of iodine are heated in a small test-tube, and the iodine 

 vapour allowed to come in contact with them by tilting the test- 

 tube. By this method the vitally stained mitochondria lose 

 their colour, becoming dark brown in appearance, but are well 

 preserved temporarily. 



764. Staining of Mitochondria with Other Basic Dyes. 

 LuDFORD {Arch. f. exjper. Zellforsch., xvii, 1935, p. 339) has 

 pointed out that mitochondria can be well stained with methylene 

 blue, especially in cells of tissue cultures. This staining can also 

 be temporarily fixed by the iodine vapour method of Lewis (see 

 above). For vitally staining cultures proceed as described above 

 for staining with Janus green, employing methylene blue in a 

 dilution of 1 in 10,000 in Ringer solution for four to eight minutes. 

 For intra-vitam staining, inject a 0-5 per cent, solution in Ringer 

 solution, and kill animals after fifteen to thirty minutes. For 

 supravital staining, incubate teased tissues in a 1 in 10,000 dilution 

 in Ringer solution, for fifteen to thirty minutes. 



Mitochondria vitally stained with this dye are rapidly de- 

 colourised on exposure to bright illumination, which also inhibits 

 the staining process. 



Other basic dyes which have been found to stain mitochondria 

 of some cells are toluidine blue and brilliant cresyl blue. 



765. Vital Staining and the Golgi Apparatus.* No dyestuff has 

 yet been discovered to stain the Golgi apparatus specifically. Most 

 investigators have not found it to stain with the dvestuffs usuallv 

 employed for vital staining. However, staining of the Golgi 

 bodies in invertebrate germ cells has been reported. This litera- 

 ture was reviewed by Bowen {Anat. Rec, xxxviii, 1928, p. 293). 

 In fibroblasts growing in tissue cultures Ludford {loc. cit., 1935) 



* Neutral red has been claimed to stain the dictyosomes by Perrincito, 

 Grabowska, Tuzet, and Wilson in snail, crayfish and scorpion. These 

 appear to be the only cases in the literature (see J. Morph., 1937). — Eds, 



