TREPONEMA PALLIDUM 



252 



TRICHINELLA SPIRALIS 



in fresh autopsy tissues. This is 

 Krajian's modification of Dieterle's 

 method (Am. J. Syphilis, 1933, 17, 127) 

 as amplified in Stain Techn., 1935, 10, 

 68. Fix tissue 5 mm. thick 10 min. in 

 10% formalin, 70°C. Cut frozen sec- 

 tions 5-7 microns. Place in 2% aq. 

 sodium cobalti nitrite 5 min. Wash 2 

 changes aq. dest. Mordant for 15 min. 

 at 70°C. in uranium nitrate 1 gm. ; 85% 

 formic acid, 3 cc; glycerin, 5 cc; 

 acetone, 10 cc. ; 95% alcohol, 10 cc. 

 Wash quickly in aq. dest. Develop 

 5 min. in 10 cc. of following mixture + 

 1 drop albumin-glycerin before use 

 (hydroquinone, 0.62 gm. ; sodium sulfite, 

 0.12 gm. ; acetone, 5 cc. ; 40% neutral 

 formaldehyde, 5 cc; pyridine, 5cc.; 

 sat. gum mastic in 95% alcohol, 5 cc, 

 aq. dest., 30 cc). Wash few sec. aq. 

 dest. Then warm silver solution 15-25 

 sec and wash in aq. dest. Keep all 

 solutions in cool place. (Original gives 

 treatment with 0.75% aq. silver nitrate 

 at 70°C. for 1 hr. upon the development 

 in hydroquinone mixture.) 



7. Levaditi's block silver method 

 (Mallory, p. 293). Fix tissue pieces 

 (1 mm. thick) in 10% formalin, 24 hrs. 

 Rinse in aq. dest. 95% alcohol, 24 hrs. 

 Transfer to aq. dest. and leave until 

 tissue sinks to bottom. Fresh 1.5-3% 

 aq. silver nitrate at 37 °C. in dark 3-5 

 days changing 3 times. (The stronger 

 silver is advised for tissues excised 

 during life.) Wash in aq. dest. Re- 

 duce 24-72 hrs. in dark at room tempera- 

 ture in : aq. dest., 100 cc; formalin, 

 5 cc ; pyrogallic acid, 2-4 gms. Wash 

 in aq. dest. Dehydrate in 80, 95 and 

 absolute alcohol. Clear in oil of cedar 

 wood, imbed in paraffin, mount 5m sec- 

 tions on slides, remove paraffin and 

 mount in balsam. Treponemas, black. 



8. Heitzman's modification of the 

 Warthin-Starry and Nieto's methods as 

 given by Mallory (p. 293). Cut frozen 

 sections 15^ or less of 10% formalin fixed 

 tissue. Place directly in pyridine, 

 10 min. Wash in aq. dest., 3 changes. 

 1% aq. uranium nitrate at 37°C., 15 

 min. Wash quickly in aq. dest., 2 

 changes. 0.25% aq. silver nitrate at 

 56°C., 15-30 min. Develop until dark 

 brown in following mixture made im- 

 mediately beforehand by pipetting into 

 a beaker: (1) 15 cc. 5% aq. gelatin at 

 56°C.; (2) 3 cc. 2% aq. silver nitrate; 

 (3) 0.5 cc. 1% aq. hydroquinone. Re- 

 move and thoroughly wash in warm aq. 

 dest. Dehydrate on slide adding by 

 pipette increasing alcohols to absolute. 

 Clear in benzol and mount in balsam. 

 A heavy black ppt. indicates too long 

 development. Treponenmas, black. 

 See Warthin-Starry method. 



9. For routine paraffin sections, 

 Steiner, G., J. Lab. & Clin. Med., 1939, 

 25, 204-210. Fix in 10%, formalin and 

 make sections 9-10 microns. Remove 

 paraffin with xylol. Pass through 2 

 changes abs. ale Treat \-\^ min. in 

 4% uranium nitrate in abs. ale, 20 cc. ; 

 25% gum mastic in abs. ale, 40-50 cc. ; 

 abs. ale, 20-30 cc. Wash in at least 3 

 changes aq. dest. until streaks of gum 

 mastic are removed. 0.1% aq. silver 

 nitrate in water bath at 100°C., 1-1| 

 hrs. Wash in aq. dest. Then through 

 80% and 95% to abs. ale 10-12.5%, gum 

 mastic in abs. ale 5 min. Repeat 3 

 washings described in aq. dest. Re- 

 duce 20-30 min. in: hydroquinone, 10 

 gm.; 12.5% gum mastic in abs. ale, 

 1 cc. ; aq. dest., 200 cc (with tempera- 

 ture gradually raised to 100°C.). Wash 

 thoroughly in aq. dest. Counterstain 

 with hematoxylin and eosin if desired. 

 Dehydrate in abs. Clear in xylol and 

 mount in balsam. The advantages are 

 speed and decrease in confusing silver 

 deposits. See Steiner's illustrations. 

 A technician's experience with Steiner's 

 method has been published (Wilson, 

 R. A. J., Am. J. Clin. Path., 1946, 16, 

 21-24). 



10. Nigrosine has been proposed as a 

 negative stain for treponema (Dienst, 

 R. B. and Sanderson, E. S., Am. J. 

 Public Health, 1936, 26, 910). Com- 

 parison of dark field, nigrosine stain 

 and Kahn test in diagnosis (Nagle, N., 

 J. Lab. & Clin. Med., 1939-40, 25, 660- 

 661). 



11. Ziehl's fuchsin stain (Perrin, T. 

 G., Am. J. Clin. Path., 1943, Tech. 

 Suppl., 7, 28). Make smears on slides 

 of exudate secured by compressing base 

 of chancre or by scraping surface of 

 ulcer. Dry in air and fix by heat in 

 flame, if desired. Stain 2 min. while 

 heating, or for 6 min. at room tempera- 

 ture, being careful not to let the stain 

 dry. The stain is aq. dest., 10 cc; 

 commercial formalin, 1 cc; acetic acid, 

 1 cc; Ziehl's fuchsin (Ziehl's Carbol- 

 Fuchsin) 4 cc. Wash in water, moving 

 gently, and dry in air. 



Triacid Blood Stain, see Ehrlich's. 



Tri-Amino Tri-Phenyl Methane Dyes. 

 These are the rosanilins. Examples: 

 acid fuchsin, acid violet, anilin blue WS, 

 basic fuchsin, benzyl violet, crystal 

 violet, ethyl green, ethyl violet, Hof- 

 mann's violet, iodine green, isamine 

 blue, magenta II, methyl blue, methyl 

 green, methyl violet, new fuchsin (ma- 

 genta III), pararosanilin (magenta O), 

 rosanilin (magenta I), spirit blue, vic- 

 toria blue B and R and victoria blue 4R. 



Trichinella Spiralis. Mallory (p. 304) gives 

 as a rapid method of diagnosis the 



