STAINS 



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STAINS 



ric acid and heat until it boils ; then place the 

 cover glass in Klug's fluid (pepsin, o. 1%, and hydro- 

 chloric acid, 0.55^); after 3 or 4 minutes wash in 

 water, dry, fix in the flame, cover with carbol-fuchsin, 

 and heat until it steams; when cool, decolor in 4 .5 

 sulfuric acid, wash in water, and counterstain with 

 malachite green for 2 or 3 minutes. Apathy's Gum- 

 syrup, dissolve 50 gm. each of picked gum arabic, 

 cane-sugar uncandiedi, and distilled water over a 

 water-bath and add 0.05 gm. of thymol. Apathy's 

 Hematein. (a 1 Alum 9 parts, glacial acetic acid 3 

 parts, salicylic acid O.I part, water iod parts. (£) 

 Hematoxylin I part, 70 % alcohol 100 parts, and pre- 

 serve for six weeks. Mix equal parts of a and b and 

 glycerol. This is the hematein mixture I A. Apa- 

 thy's Method. I. For neurofibrils : Fix the tissue in a 

 sublimate fluid and preserve in 90^ alcohol. Stain in 

 bulk for 48 hours in Apathy's hematein mixture I A; 

 wash up to 24 hours in absolutely pure distilled water, 

 transfer to spring- water, and after from 3 to 5 hours re- 

 turn to distilled water for 2 hours ; dehydrate rapidly in 

 absolute alcohol and embed in paraffin, celloidin, orgly- 

 cerin jelly. Protect from light while in chloroform or cel- 

 loidin. Mount in a resin or in neutral glycerol. 2. For 

 nerve-endings: ForegilJing. The tissue must be so 

 thin that light can stream through it, hence a membrane 

 or section. Place in 1 '' e gold chlorid solution for several 

 hours, in the dark ; transfer to I % formic acid and ex- 

 pose for from 6 to 8 hours to diffuse daylight in sum- 

 mer, to direct sunlight in winter. The temperature of 

 the acid must not be allowed to rise above 20 C. 

 Aftergilding. Fix the tissue in sublimate or in a mix- 

 ture of equal parts of saturated solution of sublimate 

 in 0.5^ salt sol ution and I a c osmic acid. Embed in 

 paraffin or celloidin, fix the sections to slides, and 

 treat them with iodin-alcohol. Place in the gold bath 

 1 or weaker) overnight, rinse in water or dry with 

 filter-paper, and reduce in formic acid, as in foregild- 

 ing. Stand the slides on end in a slanting position, 

 with the sections looking downward. The sections 

 may be counterstained and mounted in any medium. 

 3. For objects saturated with water: Infiltrate with 

 thin glycerin-gelatin ; evaporate in a desiccator kept at 

 the melting temperature of the mass. Embed in a 

 tray and harden and cut in absolute alcohol. Apa- 

 thy's Mixture. I. Equal parts of I ^ osmic acid and 

 saturated solution of mercuric chlorid in 0.5% salt so- 

 lution. 2. Mercuric chiorid, 3 gm.; sodium chlorid, 

 0.5 gm.; 50^ alcohol, 100 c.c. Argutinsky's 

 Method for attaching celloidin sections to the slide: 

 Spread a thin layer of albumin fixative on the slide; 

 warm it ; on this arrange the sections, and keep them 

 moist with 70^ alcohol; absorb the alcohol with filter- 

 paper, cover the sections with 8 or IO layers of filter- 

 paper, and with the finger on the paper press the sec- 

 tions on to the slide. Stain at once or preserve in 

 distilled water or- 70^ alcohol. Arnold's Method. 

 I. Sterilize a thin microtome section of elder-pith in 

 boiling 0.6^ salt solution; place it on a cover-glass 

 (the edges of which are coated with vaselin), charge 

 it with a drop of blood, and place the preparation on 

 a slide with a ground cell. The blood on the section 

 of elder-pith can be fixed with any of the usual re- 

 agents or films on slides can be prepared in the usual 

 way. 2. Fix blood in any suitable medium, spread it 

 on a plate and let it drv ; then pass over it a thin 

 layer of thin celloidin, drain off the excess and let it 

 dry. The celloidin with the blood can then be 

 stripped off as a thin membrane and stained. Arn- 

 stein's Method for tactile corpuscles: Macerate 

 pieces of skin for 24 hours in lime-water; remove the 

 horny stratum and treat for 5 minutes with 0. 25 <f so- 



lution of gold chlorid ; place for 24 hours in distilled 

 water; the precipitate formed is removed by putting 

 the skin in a 0.25 °c solution of potassium cyanid 

 and brushing with a camel' s-hair pencil. Mount in 

 balsam. Aronson-Phillipp Mixture, for staining 

 the granules of leukocytes : Prepare saturated aqueous 

 solutions of orange G, acid-rubin extra, and crystalline 

 methyl-green; clear by sedimentation. Mix 55 c.c. 

 of orange G, 50 c.c. of acid rubin, loo c.c. of dis- 

 tilled water, and 50 c.c. of alcohol ; to this mixture 

 add 65 c.c. of methyl-green plus 50 c.c. of distilled 

 water and 12 c.c. of alcohol. Let the solution stand 

 several weeks before using. Ascites-agar. See 

 Kiefer' s Medium and Kanthack' s Medium. Asshe- 

 ton's Method for mammalian embryos less than 10 

 days old : From I to 3 hours after the death of the 

 animal inject into the upper end of the uterus enough 

 of 0. 25 f c to o 5 f c solution of chromic acid to distend 

 the organ and smooth out the folds of the mucosas so 

 that the ova will float free in the liquid. Ligate the 

 lower end of the uterus and place it for 2 days in 

 0.5 ' ( chromic acid. Empty the contents in a watch- 

 glass and search for the ova with the microscope. 

 Stain in toto with carmine or hematoxylin and embed 

 in paraffin. Auburtin's Method for celloidin sec- 

 tions : Transfer the sections from the knife to the slide 

 and arrange before the alcohol eyaporates. Press over 

 them a strip of filter-paper and before the sections are 

 quite dry pour over them carefully several times a mix- 

 ture of equal parts of absolute alcohol and ether. 

 When the alcohol-ether has evaporated, the sections 

 will be fastened by a thin even membrane of adherent 

 celloidin. Azoulay's Method for medullated nerve- 

 fibers: Harden in Muller's fluid and embed in cel- 

 loidin. Place the sections for 5 minutes in osmic acid 

 solution ( I : 500 or iooo), wash in water and transfer 

 into 5% or io^p tannin solution and heat for from 2 to 

 5 minutes or until it steams ; wash ; stain with carmine 

 or eosin, and mount in balsam. The medullary 

 sheaths are gray to blue-black. Bacterial Suspen- 

 sions for testing disinfectants : Mix fresh cultures 

 from 3 or 4 tubes with 10 c.c. of sterilized distilled 

 water ; filter through glasswool and place in a water- 

 bath at 37.5° C. and frequently agitate, until on micro- 

 scopic examination bacteria in clusters cannot be 

 detected. Transfer 3 c.c. each into several sterilized 

 test-tubes and add an equal volume of the germicide, 

 of double the strength to be tested. At intervals of 2, 

 5, 10. 20, 30, and 60 minutes inoculate bouillon or 

 agar tubes and put them in the incubator for one week. 

 Balzer's Method for the demonstration of dermato- 

 phytes: Treat the fungus and attached scales and 

 hairs with alcohol and ether; stain for a few seconds 

 in alcoholic solution of eosin; dehydrate, clear, and 

 mount in balsam. Barfurth's Method for the egg- 

 cells of amphibia : Fix the eggs in water heated to 

 8o° C. or in chromic-acetic acid heated to the same 

 degree. For the removal of the envelopes treat with 

 javelle water diluted threefold. Eggs fixed in hot 

 water may be preserved in their envelopes in a mix- 

 ture of alcohol 125 parts, glycerol 25 parts, water 

 350 parts. Barker's Method for the detection of 

 iron in the granules of eosinophil leukocytes: Heat 

 a cover-glass film on a copper bar at 120 C. for I or 

 2 hours. Put a drop of a fresh solution of ammonium 

 sulfid on the film and immediately place the cover on 

 a slide with a drop of glycerol, so that the latter and 

 the sulfid will mix. Put the preparation in the oven 

 at 6o° C. After from 24 to 48 hours the yellow- 

 green iron reaction of the granules and the greenish- 

 black reaction of the nuclei of the eosinophil leuko- 

 cytes can be seen. Baumgarten and Jacoby's 



