382 



METHODS AND FOEMULAS 



DS 21.11 



Sections prepared in this manner are 

 permanent, and the process is so simple 

 that it can be most warmly recommended 

 as an introduction to plant section-stain- 

 ing techniques for an elementary class. 

 The sections are, however, clearly enough 

 differentiated to be used for instructing a 

 class at any rank, and they will generally 

 be found much better for this purpose 

 than complex quadruple-stained sections 

 in which the cytological detail all too 

 often tends to obscure the clarity of 

 morphological detail, which is the chief 

 requirement in this type of teaching. 



process of differentiation is also one of 

 dehydration, hence the sections may now 

 be removed with a section lifter from the 

 differentiating solution and placed in a 

 clearing agent. The writer's preference in 

 this method is for terpineol, which has 

 all the advantages of clove oil without the 

 disadvantage of tending to make the sec- 

 tions brittle so that they crack on mount- 

 ing. All the sections may be passed 

 through the differentiating solution and 

 accumulated in terpineol, where they may 

 remain until removed to a slide. Here they 

 are covered with balsam and a coverslip 

 is added. 



21.11 BONE AND CALCIFIED TISSUES 



The techniques in the present section and in the section immediately following, com- 

 prise a rather miscellaneous group, for they include both the methods intended for the 

 staining of skeletons in wholemounts, and methods developed differentiallj^ to stain 

 skeletal structures in serial sections intended for reconstruction. 



None of these methods need be employed in those cases in which it is desired to pro- 

 vide a good general stain of a section, the other tissues of which should appear in con- 

 trast. Most of the methods in sections DS 12.3, DS 13.2, and DS 13.4 in the last chapter 

 give clear differentiation of bone from cartilage in general sections. The writer's choice 

 is for the technique of Patay 1934 (Chapter 20 — DS 12.32) in which decalcified bone is 

 stained a brilliant green in contrast to the blue of the cartilage. 



21.11 Bechtol 1948 20540b, 23:3 



REAGENTS REQUIRED: A. 0.01% Biebrich scarlet; B. water 100, citric acid 2.1, methylene 



blue 0.001; C. 2.1% citric acid 

 method: [formaldehyde-fixed and hydrogen-peroxide-bleached, specimens]-^ water—* 

 A, 24 hrs. — * 95% ale, till no more color comes away — > B, 24 hrs. -^ C, till differenti- 

 ated — > balsam, via usual reagents 

 recommended for: differentiation of bone (red) and cartilage (blue) in wholemounts. 



21.11 Bock 1924a 23632, 40:318 



reagents required: A. DS 11.121 Hansen 1905; B. glycerol 50, acetic acid 50; C. 1% 



eosin B in 95% ale. 

 method: [sections of material fixed in F 7000.1000 Bock 1924 and decalcified in AF 21.1 



von Ebner (1891)] — > water -> A, 12-18 hrs. -^ thorough wash — > B, till bone alone 



remains stained, 5-30 mins. -^ running water, 1 hr. -^ C, 32^2 mins. — > balsam, via 



usual reagents 



21.11 Bock 1924b 23632, 40:318 



REAGENTS REQUIRED: A. F 7000.1000 Bock 1924; B. AF 21.1 von Ebner 1890; C. 5% 

 potassium alum; D. DS 11.124 Hansen 1905; E. glycerol 50, acetic acid 50; F. 0.4% 

 eosin Y in 95% ale. 

 method: [fresh tissues] —>■ A, 1 wk. to 1 month -* B, till decalcified — > rinse -^ C, 24 hrs. 

 -^ running water, 1-2 days — ♦ [10 fi celloidin sections] — > D, 18 hrs. -^ E, till differen- 

 tiated 5-20 mins. -^ running water 1 hr. -^ F, 5 mins. -^95% of ale, wash-> bal- 

 sam, via usual reagents 

 recommended for: demonstration of osteogenesis in decalcified tissues. 



21.11 Cretin 1937 4285a, 14:163 



REAGENTS REQUIRED: A. ADS 12.2 Cretin 1937; B. water 100, hematoxylin 1.2, alizarin 



red S 6, phosphomolybic acid 0.04 

 method: [sections of decalcified bone fixed in F 5000.1020 Cretin 1937] -^^ water -> A, 



36 hrs. -^ wash -» B, 1-24 hrs. -^ wash -> balsam, via usual reagents 



