CHONDRIOSOMES 



73 



CHOR'S MODIFICATION 



that the fatty acids could then be 

 visualized. However, hydrolysis of 

 such substrates is slow and probably 

 caused by a non-specific esterase. 

 Koelle, G. B. and J. S. Friedenwald 

 (Proc. Soc. Exp. Biol, and Med., 1949, 

 70, 617-622) proposed the use of acetyl- 

 thiocholine iodide as a substrate, and 

 obtained beautiful localizations within 

 the central nervous system. These au- 

 thors have also differentiated between 

 the true and pseudocholinesterases by 

 means of specific inhibitors (J. Nat. 

 Cancer Inst., 1950, 10, 1364). Seligman, 

 A. M., M. M. Nachlas, L. H. Man- 

 heimer, O. M. Friedman and G. Wolf 

 (Ann. Surg., 1949, 130, 333-341) have 

 proposed a naphthyl derivative of ace- 

 tylcholine as a substrate. After hydro- 

 lysis, the naphthyl moiety is visualized 

 by diazotization. 



Chondriosomes, see Mitochondria. 



Chondriotin Sulphuric Acid. Present in 

 cartilage and bone, stains metachro- 

 matically with basic dyes, described in 

 detail by Lison, L., Arch, de biol., 

 1935, 46, 599-668. See Muccproteins. 



Chorioallantoic Membrane. 1. Vital stain- 

 ing of virus lesions in membrane (Cooke, 

 J. V. and Blattner, R. J., Proc. Soc. 

 Exp. Biol. & Med., 1940, 43,255-256). 

 Place 1 cc. 0.5% aq. trypan blue directly 

 on membrane through window in shell. 

 Rotate egg gently and return to incu- 

 bator, 10-30 min. Small lesions require 

 longer time to stain than large ones. 

 Remove membrane, wash it gently in 

 physiological saline and fix flat in 10% 

 formalin, a few minutes. Make up 

 glycerin jelly by soaking 5 gms. gelatin 

 in 44 cc. aq. dest. Then add 50 cc. 

 glycerin and 1 cc. phenol. Heat gently 

 and stir. Flatten membrane on a 

 2 X 2.5 in. slide, warm glycerin jelly to 

 about 70°C. Add drop by drop to mem- 

 brane until well covered. Flame a cover 

 glass and apply with slight pressure until 

 it has begun to set. Remove hardened 

 jelly around edges and seal with balsam. 

 Foci of virus increase are sharply marked 

 by clumps of deep blue stained cells. 



2. Cultivation of microorganisms. 

 The membrane has been shown to be an 

 excellent medium for the cultivation of 

 viruses by Goodpasture, E. W., Wood- 

 ruff, A. M. and Buddingh, G. J., Am. J. 

 Path., 1932, 8, 271-282 and many others. 

 Its usefulness has been extended to 

 Rickettsiae and spirochetes by Good- 

 pasture, E. W., Am. J. Hyg., 1938, 28, 

 111-119, to fungi by Moore, M., Am. 

 J. Path., 1941, 17, 103-125 and to acid- 

 fast bacteria by Moore , M . , Am . J . Path . , 

 1942, 18, 827-847. This method of inocu- 

 lation has the advantage over laboratory 

 animal inoculation in that lesions will 



develop in the former within 5-8 days as 

 compared to weeks or months in the 

 latter; most organisms will produce defi- 

 nite and usually characteristic lesions in 

 the chick membrane, whereas they may 

 have no effect on experimental animals, 

 often requiring human subjects; and 

 because the lesions are so readily visible 

 and traceable the chlorioallantois serves 

 well as a means of virulence deter- 

 mination. 



The technique is essentially that of 

 Goodpasture and Buddingh (E. W.and 

 G. J., Am. J. Hyg., 1935, 21, 319-360) 

 with some slight changes. Fertile eggs 

 are incubated 12 days in an electrical 

 thermostat-controlled incubator regu- 

 lated to maintain a temperature of 98°F. 

 The eggs are turned twice daily. A cm. 

 square window is cut in the shell above 

 the embryo, exposing the chorioallantoic 

 membrane. The position of the em- 

 bryo is determined by candling. The 

 membrane is then inoculated directly 

 with the fungus and the window is cov- 

 ered with a sterile coverslip and sealed 

 with a paraffin-vaseline mixture (9 parts 

 vaseline, 1 part paraffin). After inocu- 

 lation, the eggs are set in a bacteriologic 

 incubator and maintained at a tempera- 

 ture of approximately 33°C., without 

 turning. The membrane is watched 

 daily through the window. When the 

 inoculated area has shown marked 

 change, the shell is cut below the window 

 and the membrane exposed. The 

 chorioallantois is cut with a pair of fine 

 curved-end scissors, removed, fixed in 

 Zenker's solution (with 5% glacial 

 acetic). After washing, dehydrating, 

 clearing in xylol, and imbedding in 

 paraffin, it is sectioned and stained. 

 Various staining techniques can be used 

 depending on the organism inoculated. 

 In general, for fungi, Loeffler's meth- 

 ylene blue and eosin have given satis- 

 factory results. For experimental tech- 

 nique of growing mouse sarcoma in 

 chorio-allantoic membrane, see Jacoby, 

 F. , McDonald, S. and Woodhouse.D. L., 

 J. Path, and Bact., 1943, 55, 409-417. 

 Chor's Modification of Ranson's pyridine 

 silver method was worked out in our 

 laboratory to show alterations in motor 

 end plates in biceps and triceps of mon- 

 keys in experimental poliomyelitis 

 (Chor, H., Arch. Neurol. & Psychiat., 

 1933, 29, 344-357). Fix in 1% ammonia 

 water (28% Merck) in 95% alcohol for 

 24 hrs. Wash in aq. dest., ^ hr. Pyri- 

 dine, 48 hrs. Wash in 8 changes aq. 

 dest. during 24 hrs. 2% aq. silver nitrate 

 in dark at room temperature, 72 hrs. 

 Reduce 6-8 hrs. or over night in : pyro- 

 gallic acid, 4 gm.; aq. dest., 95 cc; 

 formalin, 5 cc. Dip in water and trans- 



