DS 13.21 



DYE STAINS OF GENERAL APPLICATION 



351 



incredibly complex methods as those of 

 Gausen 1929 and Iloucke 1928. Complejv 

 methods of this type have proved of some 

 value in the investigation of pathology, 

 but are not to be recommended to the 

 routine worker. 



The combination of the tliiazin-eosin- 

 ates with other dyes than orange G (DS 

 13.22) have not been very successful, the 

 main exception being the technique of 



KuU 1914 which, though originally de- 

 signed for the demonstration of mitochon- 

 dria, is one r)f the best and surest triple 

 staining methods yet developed. The 

 formula of Rhamy 1930 is of great interest 

 in the staining of blood smears, and yields 

 pictures which are not only of as great 

 diagnostic value but also far more readily 

 preserved than are the standard eosin- 

 methylene blue mixtures. 



13.21 IN COMHIXATIUN WITH UKAXGE G 



13.21 Arnold 1909 methylene blue-safranin-orange G 1820, 3 :434 



REAGENTS REQUIRED: .-1. ADS 12.2 Lugol (1905); B. sat. sol. safranin in 70% ale; C. 



water 100, methylene blue 7, sodium carbonate 0.5; 1% orange G 

 method: [sections of chromic or dicliromate material] — > A, 5 mins. -^ wash -^ B, 4 hrs. 



— > wash ^ C, 4 hrs. -^ abs. ale, till dehydrated —>■ D, till differentiated -^ balsam, 



via usual reagents 

 result: nucleoli, ccntrosomes, red; many cell inclusions, blue; other structures, orange 



13.21 Cowdry 1943 phloxine-orange G-azur A Cowdry 1943, 69 



REAGENTS REQUIRED: .4. Water 100, phloxine 0.12, orange G 0.3; B. 0.1% azur A 



method: As Dominici 1902 below 



note: See also note on Cowdry 1943 modification under Dominici 1902. 



13.21 Dominici 1902 eosin Y-orange G-toluidine blue 6630,54:221 



REAGENTS REQUIRED: A. water 100, eosin Y 0.5, orange G 0.5; B. 0.5% toluidine blue 

 method: [water] -^^, 5-10 mins. -^ water, quick rinse —> B, 20-30 sees. -^ water. 



quick rinse -^ 95% ale, till differentiated -^ balsam, via xylene 

 note: Cowdry 1943 p. 69 recommends the substitution of 0.5% acid fuchsin for the 



eosin Y in .4 al)ove. See also Cowdry 1943 above and Mann 1894 below. 



13.21 Gausen 1929 methylene blue-orange G-magenta 66.30,97:1658 



FOR.MULA of methylene BLUE SOLUTIONS: 80% alc. 150, lactic acid 3, methylene blue 



2.5 

 FORMULA OF ORANGE G SOLUTION: 80% alc. 100, lactic acid 2, orange G 2 

 PREPARATION OF COMPLEX: Add bluc to Grange. Heat to 80°C. Cool. Filter. Save both 



ppt. and filtrate. 

 PURIFICATION OF PRECIPITATE: DLssolvc ppt. from above in 30 80% alc. Heat to 80°C. 



Cool. Filter. Save filtrate. Reject ppt. 

 PREPARATION OF WORKING SOLUTION: Mix both filtrates with 50 methylene blue solution. 



Filter. 

 REAGENTS REQUIRED: A. DS 11.43 Zichl 1890; B. working solution 

 method: [sections] — > water —> A, 5 mins. — > water, thorough wash — > B, 4 mins. — > abs. 



alc, till differentiated -^ neutral mountant, via usual reagents 

 result: nuclei, red; cartilage, blue; muscle, yellow; bone, green. 



13.21 Houcke 1928 toluidine blue-orange G-thionin-eosin-azur II 



6630, 99 :784 



PREPARATION OP STOCK SOLUTIONS: I. Mix 10 1% toluidine blue with 5 1% orange G. 

 Dilute to 100. Leave 24 hours. Decant and leave ppt. dry. Dissolve dried ppt. in 10 

 methanol. II. Add 17 1% eosin Y to 100 sat. sol. (circ. 25%) thionine. Leave 24 hours. 

 Decant. Dry ppt. Prepare 0.5% solution ppt. in methanol. III. Add 111% eosin B to 

 10 1% methylene blue. Thence as in II. IV. Add 1.25 1% eosin Y to 40 0.08% azur- 

 eosin. Filter. Dry ppt. on filter paper. Cut in strips and extract with 10 methanol. 

 V. Add 8 1% eosin Y to 10 1% toluidine blue. 



WORKING solution: water 100, stock I 0.5, stock II 1.5, stock III 1.5, stock IV 1.5, 

 stock V 1.5, 0.1% acetic acid 1.0 



