GOLGI APPARATUS 



145 



GOLGI APPARATUS 



Osmic Acid Technique: 



1. Mann-Kopsch method (Lee's Mi- 

 crotomists Vade Mecum, Uth Ed. 

 1950, edited by J. Bronte Gatenby and 

 H. W. Beams). Fix in Mann's fluid 

 (equal parts of 1% osmic acid and sat. 

 sublimate in normal saline), for J-3 hrs. 

 Wash in aq. dest. 15-30 min. 2% osmic 

 acid room temperature 10-14 days. 

 Wash in ruiming water 3 hrs. or more. 

 Dehydrate clear and embed. In the 

 sections Gatenby was able to extract the 

 blackening step "by step with turpentine 

 and thus to improve considerably the 

 preparations. 



2. Liidford's modification (Ludford, 

 R. J., J. Roy. Micr. Soc, 1926, 107). 

 Fix mammalian and avian tissues in 

 Mann's corrosive osmic solution 18 hrs. 

 Wash aq. dest. 30 min., 2% osmic at 

 30°C. for 3 days. Water at 30°C. for 1 

 day, dehydrate, clear, embed in paraffin. 

 A useful variant is to fix in the same 

 way, and wash in aq. dest. Then 

 osmicate at 35°C. for 3 daj'S, first day in 

 2% osmic, second in 1% and third in 

 0.5%. Leave in water for 1 day at 

 35°C. He recommends various counter 

 stains. See Lee's 11th Ed. (pp. 404- 

 410) for a discussion of osmication 

 methods; also Owens, H. B. and 

 Bensley, R. R. (Anat. Rec, 1929, 44, 

 79) for a careful study of factors in- 

 fluencing the osmic acid changes and for 

 their ferric chloride osmic method. 



3. Sjovall's method (Sjovall, E., Anat. 

 Hefte, 1906, 30, 261-291). Fix in 10% 

 formalin 8 hrs. Wash in aq. dest. 2% 

 osmic acid at 35°C., 2 days. Dehy- 

 drate, clear and embed. 



4. Kolatchew's method (Nassonov, D. 

 N., Arch. f. Mikr. Anat., 1924, 103, 

 437). Fix in 3% aq. potassium bi- 

 chromate, 10 cc; 1% chromic acid 10 

 cc; and 2% osmic acid, 5 cc. for 24 hrs. 

 Wash in running water 24 hrs. 2% osmic 

 acid 40°C. 8 hrs. 3-5 days at 35°C. 

 Wash in aq. dest., dehydrate, clear and 

 embed. Osmic methods sometimes im- 

 pregnate mitochondria as well as the 

 Golgi material particularly if the period 

 in osmic acid is prolonged, care must be 

 taken therefore in interpreting results. 



Silver Nitrate Methods: Tissues from 

 young animals usually respond best to 

 silver methods but ganglia from older 

 animals respond very well. 



1. Aoyaina's method (Baker, J. R., 

 Cytological Technique, 3rd Ed., 1950, 

 p. 194). Fix tissues in Aoyama's fixa- 

 tive (cadmium chloride 1 gm., neutral 

 formalin 4%, 15 cc, aq. dest. 85 cc.) 

 for 4 hrs. Wash with aq. dest. and then 

 place in silver nitrate solution for 12-14 

 hrs. Wash with aq. dest. and then 



place in silver nitrate solution for 12-14 

 hrs. After 8-12 hrs. (kept in dark or in 

 diffuse light). Wash with aq. dest. and 

 place in Aoyama's reducer (Hj'dro- 

 quinone 1 gm.. Neutral Formaldehyde 

 40% 15 cc, aq. dest. 80 cc, anhydrous 

 sodium sulphite 0.15 gm.) for 5 hrs. 

 Leave for I hr. in running water and 

 then place in 50% alcohol overnight. 

 Dehydrate and embed. 



2. Cajal's method (Carleton, H. M., J. 

 Roy. Micr. Soc, 1919, 321-329). This 

 is one of the many methods devised by 

 Cajal. It is recommended for embryos 

 and young animals. Fix in uranium 

 nitrate, 1 gm., formalin 15 cc, and aq. 

 dest. 100 cc, 8-24 hrs. Wash quickly 

 in aq. dest. 1.5% aq. silver nitrate 24^8 

 hrs. Rinse in aq. dest., hydroquinone 

 2 gm., formalin 6 cc, aq. dest. 100 cc, 

 anhydrous sodium sulphite 0.15 gm. 12 

 hrs. Wash in aq. dest., dehydrate 

 quickly, clear, embed and section. 



3. Da Fano's method (Da Fano, C., J. 

 Roy. Micr. Soc, 1920, 157-161). Here 

 the uranium nitrate is replaced by co- 

 balt nitrate. In other respects the 

 technique is similar. Da Fano has, 

 however, so carefully attempted to con- 

 trol troublesome experimental condi- 

 tions that the various steps are given in 

 detail. Fix in cobalt nitrate 1 gm., aq. 

 dest. 100 cc, formalin 15 cc 6-8 hrs. 

 The formalin need not be neutralized 

 unless it is strongly acid. For embryos 

 and delicate tissues where there is dan- 

 ger of shrinkage reduce formalin to 6 

 cc. Cartilage and small pieces of 

 tissue (not more than 3 mm.) fix half 

 fixation time. Hollow organs (e.g. 

 stomach and intestine) fix 1 hr. then 

 cut into smaller pieces. Spinal cord, 

 cerebellum and cerebrum of adults fix 

 8-10 hrs. (fixation should never go 

 beyond 24 hrs.). Testicle, inject fixa- 

 tive through aorta then immerse testicle 

 in fixative. Wash quickly in aq. dest. 

 and impregnate in 15% aq. AgNOs 

 24-48 hrs. Very small fragments im- 

 pregnate in 1% AgNOs, 2% for tissues 

 containing much fat and for spinal 

 cord. Impregnation normally satis- 

 factory at room temperature. If un- 

 satisfactory at 35°-37°C. Wash rapidly 

 in aq. dest. and cut down tissues again 

 to a thickness of 2 mm. or less. Reduce 

 in Cajal's hydroquinone mixture, above 

 mentioned, 12-24 hrs. Wash in aq. 

 dest. i hr. Cut with freezing micro- 

 tome or embed in wax. Golgi ap- 

 paratus dark brown or black against a 

 yellow background. Tone sections 

 with gold to clear preparation. Pass to 

 water. Then 0.1-0.2% gold chloride 2 



