678 PLANT CHROMOSOMES 



direct to several changes of 70 per cent, alcohol, fifteen minutes in 

 each. 



Nemec omits acetic acid and uses 25 c.c. of 1 per cent, chromic 

 acid with 2 c.c. of formalin. Use fresh and fix about six hours ; 

 then pour off and replace with a fresh portion to act for a further 

 eighteen hours. Reduce the time for delicate structures and 

 organisms. It is erratic in behaviour. 



See also : Licent, C. R. Acad. Sci., Paris, clxx, 1920, p. 1518 ; 

 PuJiULA, J. Broteria Sci. Bot., xxi, 1924, p. 90 ; and Bull. Inst. 

 Catalana d'Hist. Nat., ii, 1925, p. 168 ; Langlet, Svensk. Bot. 

 Tidsk., xxi, 1928, p. 397; Delaunay, Mem. Soc. Nat. Kiev, 

 XXV, 1915, p. 64. 



1358. Dehydrating fixatives are suitable for rapid methods 

 (Heitz and McClintock), or used as a pre-treatment in conjunction 

 with an aqueous fixative following Kihara's method. 



Yasui {Cytologia, v, 1933, p. 140) finds ethyl alcohol, in con- 

 centrations from 70 per cent, to absolute, satisfactory for fixing 

 pollen mother-cell smears. It is very suitable for use with 

 Fuelgen's Nuclealfarbung. Of alcoholic combinations, Carnoy is 

 widely used. Farmer and Shove {Quar. J. Micr. Sci., xlviii, 

 1905, p. 559) have given two modifications, a weaker solution 

 (6 parts absolute alcohol to 1 part glaciaj acetic acid) and a stronger 

 solution (2 parts absolute alcohol to 1 part glacial acetic acid). 

 Carnoy and Lebrun is also satisfactory. Fixation is rapid. Wash 

 afterwards in 95 per cent, alcohol. 



Of other fixatives, Gilson, Helly modification of Zenker, 

 Hermann, Juel and Merkel may be tried if necessary. 



Some of Allen's modifications of Bouin (picro-formol-acetic), 

 particularly B-15 and B-3, are useful. Fix ten minutes to a few 

 hours, according to the material. Wash in water and grade up 

 to 70 per cent, alcohol. Hold here a few days, using frequent 

 changes of 70 per cent, alcohol to which a few drops of saturated 

 lithium carbonate solution has been added. Following Bouin 

 fixations, brilliant staining with Heidenhain's haematoxylin is 

 obtainable, but gentian violet-iodine is difficult. P.F.A,-3 (B-3 

 without chromic acid) is very useful since material can be left in 

 it for long periods without danger of over-fixation. 



Skovsted [Ann. Bot., xlvii, 1933, p. 227), working on Gossypium 

 in the tropics, recommends pre-fixation in Winge's Picro-Carnoy 

 {Zeits. Zell. Mikr. Anat., x, 1930, p. 683), followed by an osmic 

 acid modification of Allen's Bouin. Remove calyx and top of 

 corolla of buds and place five to fifteen minutes in Picro-Carnoy 

 at 37° C. 



Picro-Carnoy is : 



8 per cent, mercuric chloride in absolute 



alcohol . . . . . .1 part 



Chloroform ...... 2 parts 



