CAJAL'S 



54 



CALCIUM 



render phospho- and galactolipines less 

 soluble. Bourne (p. 106) refers to 

 Joyet-Lavergne's claim that cadmium 

 lactate reacts with glutathione in the 

 cell producing a cadmium glutathione 

 compound which is microscopically 

 visible. 



Cajal's. Properly the name should be listed 

 as Ramon y Cajal. 1. Brom-formol- 

 silver method for neuroglia. Details 

 supplied by Dr. J. L. O'Leary. Fix 

 small fresh pieces, 3-15 days, in: aq. 

 dest., 85 cc; formalin, 15 cc; ammo- 

 nium bromide, 2 gm. Cut 25/i frozen 

 sections and return to: aq. dest., 50 cc; 

 formalin, 6 cc; ammonium bromide, 

 3 gm. for 4-6 hrs. at 30-38°C. or for 8-10 

 hrs. at room temperature. Wash for a 

 few seconds in aq. dest. Place in the 

 following fluid in a porcelain dish and 

 heat over the flame: aq. dest., 10-15 cc; 

 ammoniacal silver oxide, 5 cc; pyridine 

 C.P., 4-5 drops. (To prepare silver 

 oxide solution: Take 10 cc. 10% silver 

 nitrate, add 12 drops 40% NaOH. Col- 

 lect the ppt., wash 5-6 times with aq. 

 dest., then add ppt. to a beaker con- 

 taining 60-70 cc. aq. dest. Redissolve 

 with least quantity of ammonia neces- 

 sary. If too much ammonia is added, 

 results are bad.) Remove when sec- 

 tions have reached a tobacco brown 

 color. Wash through 2 changes aq. 

 dest. not more than 5 sec. in all. Re- 

 duce in 5% formalin for 2-3 min. Tone 

 with 0.2% aq. gold chloride and fix in 

 5% aq. sodium hyposulfite. After 

 washing carry to 95% alcohol, carbol- 

 xylol, xylol balsam. See Microglia and 

 Oligodendroglia. 



2. Chloral hydrate method as de- 

 scribed by Willard, D. M., Quart. J. 

 Micr. Sci., 1935-36, 78, 475-485 for 

 innervation of adrenal. Fix for 24 hrs. 

 in : chloral hydrate, 2.5 gm. ; 95% alcohol, 

 40 cc. ; aq. dest., 40 cc. ; pyridine, 20 cc 

 Wash in aq. dest. until smell of pyridine 

 disappears. 97% alcohol, 24 hrs. Wash 

 again in aq. dest. and transfer to 2.5% 

 aq. silver nitrate at 37°C. for 9-12 days 

 (longer times better for nerve cells). 

 Wash for 1 min. in aq. dest. Reduce for 

 12-24 hrs. in: hydroquinone, 1 gm.; 

 neutral formol, 10 cc; aq. dest., 90 cc. 

 Dehydrate rapidly, embed in paraffin 

 and cut 15-30m sections. Nerve fibers, 

 black; background, yellow. 



Cajal Silver Methods. These depend 

 mainly on silver impregnations reduced 

 by photographic developers such as 

 hydroquinone. They have all been 

 very greatly improved by a preliminary 

 fixation and in other ways and have 

 played a leading r61e in neurology. See 

 Ranson pyridine method and other 

 modifications given by Addison (Mc- 



Clung, pp. 452-463). Many techniques 

 spring from a combination of Cajal and 

 Bielchowsky methods. 



Calcareous deposits. Vital staining with 

 Alizarin Red S (Ham, A. W., Arch. 

 Path., 1932, 14,613-626). 



Calciferol, see Vitamin D2. 



Calcium. There is no absolutely specific 

 microchemical test for calcium in sec- 

 tions. A critical account by Cameron 

 (G. R., J. Path, and Bact., 1930, 33, 

 929-955) affords instructive reading. 



1. von Kossa test. Sections are trans- 

 ferred from aq. dest. to 10% silver 

 nitrate and exposed to bright light for 

 30 min. or more. Wash carefully in aq. 

 dest. Mount in glycerin, or dehydrate 

 clear and mount in balsam. Inorganic 

 material in most cases calcium phos- 

 phate or carbonate is deep black. See 

 comments of Gomori, G., J. Mt. Sinai 

 Hosp., 1945, 11, 317-326. Test has been 

 adapted to investigation of bone by 

 McLean, F. C. and Bloom, W., Anat. 

 Rec, 1940, 78, 333-359. 



2. Alizarin. Sections from aq. dest. 

 are stained in 1% aqueous alizarin S 

 (sodium alizarin sulphonate) or in 1% 

 alcohol tetra-hydroscy-anthraquinon (or 

 anthrapurpurin) for an hr. or more. 

 They are then differentiated in 1 part 

 concentrated ammonia and 9 parts 

 absolute alcohol. This is followed by 

 rapid washing in acid alcohol (hydro- 

 chloric acid 5 cc, 95% alcohol 95 cc). 

 It may be desirable to alternate alkali 

 and acid alcohols 2 or 3 times. Wash 

 thoroughly in aq. dest. ; dehydrate clear 

 and mount. The alizarin forms a fast 

 compound with earthy salts especially 

 calcium more easily in young than in 

 old bones. Substances may exist in the 

 tissues that inhibit the combination 

 (see Bone, Madder staining). 



3. Hematoxylin. This is not, as is 

 generally supposed, a stain for calcium 

 though it may color calcium as well as 

 other materials when mordanted with 

 chromium salts or alum. According to 

 Cameron, in bone, "staining with 

 hematoxylin is dependent on the essen- 

 tial ground substance and the presence 

 of certain heavy metals especially iron 

 chromium and aluminum ; it has no 

 direct relation to calcium salts." He 

 thinks that areas of pathological calci- 

 fication which stain deeply with alum 

 hematoxylin do so because of the pres- 

 ence of free iron. 



4. Fluorescence x-radiation. Used for 

 thin sections of undecalcified bone. It 

 is not feasible to magnify much but the 

 method is said to be almost specific for 

 calcium (Dershem, E., Proc Nat. Acad. 

 Sci., 1939, 25, 6-10). 



5. Cretin, A., Bull. d'Hist. Appl., 



