STAINS, ETC. 



1376 



EXAMINATION OF THE BLOOD 



apart.) The outline drawing is now to be filled in 

 with the details of the drawings of the sections. This 

 is accomplished as follows : A piece of glass, of the 

 size of the intended drawing, is coated with gelatin 

 and ruled with a series of close, parallel lines with 

 differently colored inks, the colors recurring in regular 

 order. Cut the plate in two equal parts on a line per- 

 pendicular to the ruled lines. Lay one plate on the 

 outline drawing so that its cut edge covers the line cor- 

 responding to the first section to be filled in, then lay 

 the other plate on the drawing of the sections in such 

 a position that the limit of the drawing corresponds to 

 the same colored lines that cover the limits of the out- 

 line drawing. Trace on the plate that covers the draw- 

 ing of the section the outline of the internal organs. 

 Lay it against its fellow on the outline drawing, making 

 the lines correspond. Mark off the outlines of the 

 internal organs. Repeat this operation for each sec- 

 tion and connect the series of dots so made and the 

 drawing is completed. Another method of recon- 

 structing objects from microscopic sections is that 

 suggested by Born. By the aid of the camera, the 

 outlines of the sections are transferred to wax plates, 

 which are then cut out so as to correspond, in out- 

 lines as well as dimensions, to the sections equally 

 magnified in all three directions. With plates thus 

 prepared, it is only necessary to put them together in 

 the proper order to obtain a complete model. Mam- 

 malia. For the study of the early stages, the ova must 

 be obtained from the tubae (of a rabbit or other small 

 animal) several hours after copulation. Dissect out the 

 tubse and cornua of the female, allow them to cool, and 

 wait for the muscular contraction to cease. Dissect off 

 all the peritoneal investment, and slit the tubse open 

 longitudinally. The folds of the tubal mucosa are 

 spread out by means of needles and forceps, and the 

 ova searched for by means of a magnifying glass. The 

 ova are best examined in the peritoneal fluid of the 

 mother or in the aqueous humor, blood-serum, or ar- 

 tificial serum. Kolliker injects Midler's fluid or a weak 

 osmic-acid solution into the oviduct, and collects the 

 fluid that runs out in a series of watch-glasses, which 

 are examined for the ova under the microscope. Dur- 

 ing the fourth, fifth, and sixth days after copulation 

 the ova are free in the uterine cornua, and are easily 

 visible to the eye, and may be obtained in a like 

 manner as from the tubse. When the ova become fixed 

 in the uterus they are easily distinguished by the pecu- 

 liar aspect of the cornua in which there are small eleva- 

 tions at the site of each ovum. To obtain the ova it 

 is necessary to incise the cornua transversely into as 

 many segments as there are eminences, care being 

 taken to have the ova in the center of the segments. 

 The segments are then fixed to the bottom of a dis- 

 secting dish by pins, with the mesometrial surface 

 downward. Fill the dissecting dish with serum, 

 Midler's fluid, or Kleinenberg's picro-sulphuric-acid, 

 or nitric-acid solution. The ovular eminence is then 

 incised longitudinally and the ova carefully freed. 

 Preparations. To make permanent preparations of 

 the various stages of fecundation and segmentation, 

 the living ovum is placed in a I per cent, solution of 

 osmic acid, on a slide, then into Midler's or Kleinen- 

 berg's solution. In an hour the solution is changed 

 and the whole is placed in a moist chamber for two or 

 three days. It is then treated with increasing strengths 

 of glycerin, and mounted in pure glycerin, acidulated 

 with formic acid ; or ova may be stained with picro- 

 carmin after treatment with osmic acid and careful 

 washing. To demonstrate the blastoderm cells, treat 

 the living ova in a ^ per cent, solution of argentic 

 nitrate for y^ to 3 minutes, then place in distilled water 



and expose to the light. These specimens cannot be 

 rendered permanent ; they ultimately become black. 

 The blastodermic vesicle can be opened with a fine 

 needle after 3 days, and the blastoderm washed, 

 stained, and mounted in glycerin or balsam, or pre- 

 pared with gold chlorid. For embryonic areas and 

 the more advanced embryos, place ova in a 0.5 per 

 cent, osmic-acid solution until quite dark (about 1 

 hour), then treat with successive alcohols for several 

 hours. For sections, Kolliker fixes the ova in osmic 

 acid, and v. Beneden treats them for 24 hours with 1 

 per cent, chromic-acid solution, washes thoroughly, and 

 carries them through successive alcohols. Piersol re- 

 commends Kleinenberg's solution or, for young stages, 

 Altmann's 3 per cent, nitric acid. Stain small em- 

 bryos with borax-carmin or Delafield's hematoxylin 

 (see Staining Reagents), and for larger ones Henne- 

 guy's acetic acid alum-carmin gives the best results. 

 For sections, embed in paraffin and mount in balsam. 



EXAMINATION OF THE BLOOD. 

 Alkalinized Urine. Used in the enumeration of blood- 

 corpuscles. Saturate a quantity of urine with borax, 

 filter, and dilute until its sp. gr. is 1020. The contour 

 of the cells remains unchanged in this medium. 

 Auerbach's Method for Amphibian Red Blood-cor- 

 puscles. By fixing the blood-film on the cover-glass by 

 means of a saturated solution of picric acid or a mix- 

 ture of O. I to 0.25 per cent, solution of corrosive sublim- 

 ate, I per cent, solution of boric acid, I percent, sodium 

 chlorid, or 2 percent, to 10 percent, ammonium chro- 

 mate, certain differentiations of the corpuscle are pos- 

 sible. With picric-acid fixation and subsequent stain- 

 ing with eosin and anilin-blue, the cell-wall stains blue, 

 while the adjacent protoplasm within stains red. The 

 protoplasm may further be separated into a cortical and a 

 medullary layer, the former containing the hemoglobin. • 

 In picric-acid preparations the cortical layer shows a 

 beautiful network, while the medullary part is clear, I 

 like a large hole. In sublimate preparations the med- 

 ullary part has dark granules. Blood-platelets. 

 Ligate the finger and prick the pad. Wipe off the; 

 first blood that exudes, and touch the apex of the. 

 second drop with the cover-glass; drop it gently on 

 the slide, do not press it on, and platelets will have 

 their faces, not their edges, presented to view. Stir- 

 ling recommends that the finger be pricked through a 

 drop of normal saline solution containing methyl-vio- 

 let (0.75 cm. in IOOO c.c). The colorless corpuscles 

 are stained light-blue, the platelets dark-violet or dark- 

 blue. Ehrlich's Methods. Ehrlich's Granules. 

 Dry a cover-glass preparation of blood for several 

 hours at 120 C. , or rapidly over the flame of a Bun- 

 sen-burner. Stain 1 hour or longer in eosin-glycerin ; 

 wash in water, dry, and mount in balsam. ( >r, stain in 

 glycerin, 30 c.c, and 2 gm. each of aurantia, indulin, 

 and eosin. If the eosin-indulin-glycerin solution be 

 used the rf-granulations are purplish-red and the 

 nuclei bluish-black. 1. Oxyphilous, or Eosin 

 ous Granules. Cover-glass preparations of blood an 

 fixed by dry heat, as indicated, or by chemic reagents 

 corrosive sublimate, or osmic acid. The preparatiol 

 is then floated on a I per cent, aqueous soluti 

 eosin, a quarter to one minute. A trace of aceti< 

 acid added to the fluid causes the specimen to 

 stain rapidly, and the excess of dye is removed 

 all partsof the cells, except the oxyphilous granuli 

 dipping the cover-glass into a very dilute solution 

 sodium carbonate. 2. Neutrophil Granules. Thes* 

 are the e-granulations of Ehrlich. They are stainei 

 only by neutral dyes, e. t ?., acid fuchsin, fuchsin-S 

 methylene-blue. 3. Basophilous Granules. Thesi 



