CARBOL-XYLOL 



58 



CARBOWAX EMBEDDING 



Carbol-Xylol. Xylol saturated with car- 

 bolic acid crystals. After using it for 

 clearing celloidin sections, wash quickly 

 in xylol before naounting them in 

 balsam. 



Carbon from inspired air occurs abundantly 

 in lungs and bronchial lymph nodes. 

 It may be transported to the great blood 

 filters (spleen and liver) where it is 

 distinguishable by its black color and 

 by its insolubility in cone, sulphuric 

 acid which dissolves all other body 

 pigments. Fine suspensions of carbon 

 are of great service as vital stains to 

 demonstrate phagocytosis. See Hig- 

 gins' Ink and Lampblack. 



Carbon Monoxide, Measurement of, see 

 Scholander, P. F., and Roughton, F. 

 J. W., J. Biol. Chem., 1943, 148, 551- 

 563, or Glick, p. 334. 



Carbonic Anhydrase. This can be localized 

 in the oxyntic (or parietal) cells of the 

 fundus of the stomach. Davenport, 

 H. W., Am. J. Physiol., 1940, 128, 

 725-728; 129, 505-514 employed an 

 adaptation of Linderstr0m-Lang's tech- 

 nique and observed that in rats and cats 

 the parietal cells contain 5 to 6 times as 

 much of the enzyme as red blood cells 

 while the peptic cells are free from it. 

 A microspectroscopic method for demon- 

 stration of carbonic anhydrase within 

 erythrocytes depends on the action of 

 methemoglobin as an indicator which 

 changes both its color and pattern of 

 absorption spectrum with change of pH 

 from 6.5-9.5 (Keilin, D. and Mann, T., 

 Nature, 1941, 148, 493-496). For data 

 on the distribution of this enzyme in 

 lower forms, see Blaschko and Jacobson 

 (Bourne, p. 200). 



Carbonyl Compounds, water insoluble alde- 

 hydes and ketones, see critical state- 

 ment by Glick, pp. 69-72. Bennett, 

 H. S. (Am. J. Anat., 1940, 67, 151-228) 

 regarded his phenylhydrazine reaction 

 for carbonjd compounds as indicative 

 of ketosteroids in the adrenal cortex. 

 Gomori, G. (Proc. Soc. Exper. Biol. & 

 Med., 1942, 51, 133-134) however does 

 not agree unless there is additional evi- 

 dence. Glick is of the same opinion 

 that in the absence of such evidence the 

 carbonyl reactions only indicate lipid 

 aldehyde or ketone. According to 

 Albert, S. and Leblond, C. P. (Endo- 

 crinology, 1946, 39, 386-400) it is plasmal- 

 ogen instead of ketosteroid which is 

 demonstrated by the phenylhydrazine 

 reaction. 



1. Bennett's phenylhydrzine reac- 

 tion. Place frozen sections of fresh 

 tissue from microtome into M/10 ace- 

 tate buffer, pH 6.0-6.5. If sections of 

 fixed tissue are employed place in water. 

 Add 1% iodine in ale. drop by drop till 



pale straw yellow color persists. Let 

 stand 15 min. Add 1% aq. sodium 

 thiosulphate drop by drop till color is 

 lost and a small amount more is added. 

 Let stand 5 min. Wash sections re- 

 peatedly in aq. dest. Transfer sections 

 to buffered phenylhydrazine solution 

 just prepared by mixing equal volumes 

 of 2% aq. phenylhydrazine hydro- 

 chloride and the acetate buffer and by 

 removing oxygen by gently bubbling 

 through carbon dioxide for 15 min. 

 This solution is to be poured into glass- 

 stoppered bottles so that with the sec- 

 tions added no air bubbles remain under 

 the stopper. Control slides are taken 

 through the same procedures but minus 

 the phenylhydrazine treatment. 



2. Albert and Leblond's 2,4-Dinitro- 

 phenylhydrazine reaction. Saturate 

 2,4-dinitrophenylhydrazine (No. 1866 

 Eastman Kodak Co.) in 30% ale. and 

 bring pH to neutrality by addition of 

 0.2 N sodium acetate. This is the re- 

 agent. Fix tissue in formalin neutra- 

 lized with magnesium carbonate for 

 48 hrs. and wash in running water for 

 24 hrs. Cut frozen sections 10-15 n 

 and transfer to 17% ale. 4 hrs. Place 

 them in the reagent over night and 

 wash in 17% ale. 20 min. Change to 

 aq. dest. and mount in glycerol gelatin 

 (see Glychrogel). Yellow color indi- 

 cates positive reaction. 



3. Seligman, A. M. and Ashbel, R. 

 (Cancer, 1951, 4, 579-596): Frozen sec- 

 tions, 10 to 20 M, are cut from formalin- 

 fixed tissues and attached to slides by 

 air-drying, after which the formalin is 

 washed out wath several changes of 

 water. Sections are incubated for one 

 or more hours at room temperature in 

 a 0-1% solution of 3-hydroxy-2-naph- 

 thoic acid hydrazide in 50% aldehyde- 

 free alcohol and 5% acetic acid. Re- 

 move e.xcess reagent by washing for 

 2 hrs. in 50% alcohol followed by 2 hrs. 

 in several changes of water. 



A blue pigment is produced at the 

 sites of carbonyl reactivity by immer- 

 sion in a solution prepared from equal 

 volumes of absolute ale. and an aqueous 

 solution containing two parts water 

 and one part 1/15 M phosphate buffer 

 (pH 7-2), followed by the addition of 

 tetrazotized o-dianisidine in powered 

 form (50 mg. for 50 cc. of solution). 



The development of a blue color 

 reaches a maximum in one or two min. 

 The sections are washed in several 

 changes of water (acidified with a few 

 drops of acetic acid) and are mounted 

 in glycerogel. 

 Carbowax Embedding — Written by Dr. 

 H. I. Firminger, Pathology Section, 

 National Cancer Institute, Bethesda, 



