EXPERIMENTAL EQUIPMENT AND PROCEDURES 11 



Urodele Operating Medium : ( hypertoni c ) 



Stock solution 2 parte 



Great Bear Spring Water 1 part 



Urodele Growing Medium : (isotonic) 



Stock solution 2 parts 



Great Bear Spring Water k parts 



7- Locke's solution - for chick emtryos: 



NaCl 0.9 gr. 



ICCl O.OU gr. 



CaClg 0.021+ gr. (anhydrous) 



KaHCOj 0.02 gr. 



Water 100.0 cc. 



8. Special variations of the ahove solutions : 



A. Calcium- free Standard (Holtfreter' s) Solution. 



B. Sodium-free Standard ( Holtfreter ' s) Solution. 



C. Potassium- free Standard ( Holtfreter' s) Solution. 



D. Buffer-free Standard ( Holtfreter' s) Solution. 



E. Nuclear medium for germinal vesicle studies (Calcium- free Ringer's). 



NaCl 0.6 gr. 



KCl 0.01 gr. 



Glass Distilled Water 100.00 cc. 



9- Anesthetics : 



A. MS-222 (Sandoz Chem. Co., 65 Van Dam St., N.Y.C.) Excellent for embryos. 



Make up 1/30OO concentration in Spring Water and in Standard Solution of 

 the ahove concentrations. (Tricain Me thane sulfonate) Must be used fresh. 



B. Chloretone - 0.5^ in Spring Water and in Holtfreter' a. 



C. Ether - anesthetic form. 



D. Chloroform. 



E. Magnesium sulphate (Epsom salts), crystal form. 



10. Killing and fixing solutions : 



A. Smith's fixative - best for yolk-laden aicjihibian eggs. Two solutions to be 



mixed Just before use. Not to be used if discolored. 



Solution A : K Bichromate 0.5 gr. 



Water 87.5 cc. 



Solution B : Formalin 10 cc. 



Acetic (glaclal)2.5 cc. 



B. Bouin's fixative : safest of all fixatives, particularly for late stages. 



Saturated (aqueous) picric acid 75 cc. 



Formalin ( oommercial) 25 cc. 



Glacial acetic acid _ 5 cc. 



C. Bouin-Dioxan - excellent for yolk embryos and rapid technique. (See Puckett, 



1957 - Stain Tech. 12:97). Use Bouin's and Dloxan in equal parts (Dloxan 

 is volatile and toxic). 



D. Mlchaelis' fluid - For yolk embryos. 



Cone. HjjClg - aqueous 20 pts. 



Cone, picric aqueous 20 pts. 



Glacial acetic acid 1 pt. 



Water distilled 1+0 pta. 



E. KLeinberg's pi cro- sulphuric - for polar bodies and spindles (McClungJ. 



Water 200 vols. 



Cone. B^SOl^ 2 vols. 



Picric acid - to saturation 



F. Chrom- acetic fixative - excellent for cytological studies of amphibian egg 



and embryos : 



Chromic acid 10^ 25 pts. 



Glacial acetic acid 10 pts. 



Sat. aq. picric acid 100 pts. 



