STAINS 



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STAINS 



The sections will spread out in a few seconds. Drain 

 off the superfluous alcohol and arrange the sections. 

 Moisten in Sofo alcohol a strip of smooth, thick blot- 

 ting-paper, place it on the sections, and over this 

 another dry strip. Pass a smooth metal roller several 

 times over the paper and the sections will adhere to 

 the slide. Dry in the oven. They can be stained at 

 once or kept indefinitely unstained. 2. For the dem- 

 onstration of the filaments of human blood-platelets : 

 Prepare the films on cover-glass chemically clean ; dry 

 in air for 12 hours and fix in absolute alcohol ; stain 

 for 24 hours in a 1 % aqueous solution of toluidin blue, 

 wash in water, dry by means of a bellows, and mount 

 in balsam. Eosin stains the filaments red. A 

 strong solution of hemalum stains the centrosomes. 

 3. For the study of spermatogenesis in amphibia : 

 Fix the testes from 3 to 12 hours in 0.5 to 0. 1 % solu- 

 tion of osmium chlorid, wash for an hour in water, 

 treat successively with alcohol, bergamot oil, xylol, 

 again bergamot oil, and embed in paraffin. Cut 4 fi to 

 6 fi sections and stain in Benda's iron-hematoxylin, 

 adding 10% of alcohol to the dye and staining for 

 from 48 to 72 hours and differentiating in 10$ acetic 

 acid containing a very little of the iron sulfate. After- 

 stain with congo red. A triple stain can be obtained 

 by treating the sections for a few seconds with a weak 

 aqueous solution of congo red, then for 10 minutes 

 with an aqueous solution of thionin, and differentiating 

 in a very weak aqueous solution of ruthenium red. 4. 

 For the preservation of corks and paper labels : 

 When the ink is dry dip the label for a minute into 

 melted paraffin ; drain, and when the paraffin is hard 

 put the label inside the bottle with the specimen. 

 Put the new, dry cork into the melted paraffin for a few 

 minutes. Place a string on one side when inserting 

 the cork in the bottle. Pin a label to the cork and 

 then dip cork and neck of bottle into the paraffin. 

 Not only label and cork are preserved, but the alcohol 

 does not evaporate. Eosinate of Methylene-blue. 

 See Jtositi's Stains. Eosin-iodin, iodin I gm., 

 potassium iodid 2 gm., 2 c.c. of saturated solution of 

 eosin in 90^ alcohol, and 200 c.c. of distilled water. 

 Ewald's Method : Mix 3 or 4 drops of blood with 

 IO c.c. of a 0.5% solution of osmic acid in 0.6% salt 

 solution. After 24 hours siphon off the supernatant 

 liquid with Ewald's capillary siphon, add water, with- 

 draw it and add alum-carmin, and so on, finally 

 treating with $o c / c alcohol. Eyclesheimer's Method 

 for orientation of celloidin objects: I. Use metal em- 

 bedding frames with the sides and ends perforated with 

 small holes ; pass silk thread through the opjxasite 

 holes, allowing a length of about 2 inches to hang 

 loose at each end. The net of threads is made taut 

 by gluing each thread with a drop of celloidin to the 

 outside of the frame. Place the object on the net and 

 pour in the celloidin. Soak one loose end of each 

 thread in thin celloidin containing lampblack, and 

 when the mass is hard, dissolve the celloidin fastening 

 the threads to the frame by means of ether. Finally, 

 pull the threads out of the box so that the blocked 

 ends will mark the bottom of the mass and form orien- 

 tation points. 2. Arrange the sections on a slide with 

 enough alcohol to keep them moist ; cover them with 

 a strip of tissue-paper and secure it by thread passing 

 between, not over, the sections. Stain in any pre- 

 ferred way ; after clearing, cut the thread and strip off 

 the paper. Feinberg's Method for the study of the 

 structural relations of bacteria : Prepare Roman- 

 owsky-Ziemann's stain with 1.5% to 2% solution of 

 methylene-blue that has been subjected to a tem- 

 perature of 86° C. for several hours. Stain for 3 or 

 4 hours and finally for several minutes in the warmed 



fluid. Fertilization, Artificial : Shake the ripe 

 ovary of an echinoderm recently caught in a dish con- 

 taining an abundance of sea-water. The eggs appear 

 as little white dots. Remove fragment of tissue. In 

 the same way empty a ripe testicle in a separate dish 

 of sea-water ; distribute evenly in the water and add a 

 very small quantity of this sperm-containing water to 

 the water containing thg eggs. The spermatozoa pene- 

 trate the ripe eggs in 5 or IP minutes and the first 

 cleavage is complete in about an hour and a half. 

 Every 5 or 10 minutes place a large quantity of the 

 eggs in picric-acetic acid and subsequently stain in 

 borax-carmin. F., Polyspermous : Place the fresh 

 eggs of the sea-urchin for from 5 to 60 minutes in a 

 0.5% solution of chloral in sea-water. Transfer to 

 fresh sea-water and fertilize (see fertilization, Arti- 

 ficial}. Fix in picric-acetic acid and stain in borax- 

 carmin. The numerous astrospheres and the early 

 abnormal cleavage can be studied in the living object. 

 Cf. the method of Hertwig. Fich-Schultze Method 

 for the ova of amphibia : Treat the eggs in their en- 

 velopes for 24 hours with chromic-acetic acid ; remove 

 the envelopes and wash the eggs for 24 hours in run- 

 ning water ; harden in 6o^£ and Sofc alcohols, for 24 

 hours each, and stain for 24 hours in borax-carmin. 

 Embed in paraffin. Ficker's Medium for the culture 

 of tubercle bacilli : Grind the brain of a cow, calf, or 

 horse in a meat machine, add an equal volume of 

 water, and slowly heat to boiling, stirring meanwhile. 

 After cooking for 15 minutes, strain through cloth and 

 sterilize for 2 hours in steam. Add equal volumes of 

 serum and 3^ glycerin, pour into test-tubes, and co- 

 agulate in the serum oven ; or, mix with equal volumes 

 of 2.5% solution of agar and 3% glycerol, and steril- 

 ize for 2 hours by steam. Field-Martin Method 

 of celloidin-parafffn embedding : Place the thoroughly 

 dehydrated object for several hours in a mixture of 

 equal parts of absolute alcohol and toluol, then for 

 several hours in a celloidin-parafffn mixture prepared 

 as follows: dissolve celloidin in equal parts of abso- 

 lute alcohol and toluol until the mixture has the con- 

 sistence of clove-oil, then saturate with paraffin at a 

 temperature of 25 C. From this transfer the object 

 to chloroform saturated with paraffin and then to the 

 usual paraffin solution. Finotti's Method for the 

 myelin of nerves: Fix in Miiller's fluid for one 

 month. Place the sections for 10 hours in a freshly 

 prepared mixture of equal parts of I % osmic acid and 

 concentrated solution of picric acid in one-third alco- 

 hol, protecting meanwhile from the light. Fischer's 

 Method for the flagella of bacteria : Treat the 

 cover-glass films for one minute in the steaming hot 

 mordant, prepared as follows: dissolve 2 gm. of des- 

 iccated tannin in 20 c.c. of hot water and add 4 c.c. 

 of a I : 2 green ferrous sulfate solution and I c.c. of a 

 concentrated alcoholic solution of fuchsin, and filter. 

 Wash the films on water and stain in heated saturated 

 aqueous solution of fuchsin. Fisher's Eosin : Make 

 a saturated solution of water-soluble eosin (Grubler), 

 add hydrochloric acid, in slight excess, and collect the 

 precipitate on a filter; wash with water until the 

 filtrate begins to be tinged with the eosin ; let the 

 precipitate dry, powder, and for use dissolve in alco- 

 hol. Flormann's Method for coloring aclinomyces 

 in tissue sections: Stain 5 minutes in a mixture of 

 concentrated alcoholic solution of methyl-violet, 1 

 volume, 1% aqueous solution of ammonium carbon- 

 ate, 2 volumes, and water, 2 volumes : wash for 10 

 minutes in an abundance of water and treat for 5 

 minutes with 1:2: 300 iodin-potassium iodid solution ; 

 wash and extract for 20 minutes in 1 : 50 fluorescein 

 alcohol, renewed once ; wash in 95 % alcohol, treat with 



