GLYCOGEN 



109 



GOLD 



minum rubrum optimum or some 

 other good carmine 2 gm., potassium 

 carbonate 1 gm., potassium chloride 5 

 gm., aq. dest. 60 cc. Boil gently until 

 color darkens, cool and add 20 cc. con- 

 centrated ammonia. Allow to ripen 24 

 hrs. This is stock solution. Mount 

 paraffin sections, bring down to aq. 

 dest. Stain nuclei with hemato.xylin 

 as in the H. and E. technique. Transfer 

 to fresh stain (stock solution 10 cc, 15 

 cc. cone, ammonia and 30 cc. pure 

 methyl alcohol) for 20 min. Rinse in 

 3 changes methyl alcohol, dehydrate in 

 acetone, clear in toluol and mount 

 in balsam. Glycogen brilliant red. 



2. Iodine (Gage). Mount paraffin 

 sections as before, being again careful 

 to avoid unnecessary heat, and bring 

 down to water. Lugol's aq. iodine 

 10-15 min. Blot with filter paper and 

 dry in air. Mount in yellow vaseline 

 as advised by S. H. Gage (J. Comp. 

 Neur., 1917, 27, 451-465) with minimum 

 of heat. Glycogen reddish brown. 



3. Bauer-Feulgen. To make Feulgen 

 reagent dissolve 1 gm. basic fuchsin in 

 100 cc. aq. dest. by heat. Filter while 

 warm and add when cool 20 cc. normal 

 HCl. Add 1 gm. NaHSOs. Allow to 

 rest 24 hrs., when it should be of pale 

 straw yellow color. Treat deparaffinized 

 sections with 4% chromic acid for 1 hr. 

 or with 1% chromic acid over night. 

 After washing in running water 5 min., 

 place in Feulgen reagent 10-15 min. 

 Rinse 1^ min. in each of 3 changes of 

 molecular sol. NaHSOs 1 part and tap 

 water 19 parts. Wash in running 

 water 10 min. Counterstain nuclei with 

 hematoxylin if desired. Dehydrate, 

 clear and mount in balsam. Glycogen 

 deep reddish violet, nuclei lavender. 



Control. Prepare at same time some 

 sections of liver rich in glycogen. Be- 

 cause glycogen is quickly removed by 

 salivary digestion, when sample sections 

 are brought down to aq. dest., spit on 

 them and allow to rest 15-30 min. chang- 

 ing saliva several times. Wash thor- 

 oughly in water at body temperature 

 to remove mucus and stain by either of 

 the 3 above mentioned techniques. If 

 the material is then absent in such 

 sections and present in other similarly 

 stained and not digested, it is evidently 

 glycogen. Fixation by the freezing and 

 drying method is even better than with 

 the alcohol, picric, formalin mixture 

 because it is quicker and there is less 

 chance for displacement of glycogen in 

 the cells. 



See also for glycogen staining of 

 Trachoma inclusions Thygeson, P., Am. 

 J. Path., 1938, 14, 455-462. Glycogen is 

 immobilized in its natural position 



within the cells by the Freezing and 

 Drying technique (Altmann-Gersh). 

 Compare figures 3 and 4 of Bensley and 

 Gersch (R. R. and I., Anat. Rec, 1933, 

 57, 205-215) showing results by this 

 and other methods. 



A new ammoniacal silver nitrate 

 method for glycogen is described by 

 Mitchell, A. J., and Wislocki, G. B., 

 Anat. Rec, 1944, 90, 261-2C6. To pre- 

 pare silver solution dissolve 1 gm. silver 

 nitrate in 10 cc. aq. dest. and add 11 

 drops 40% aq. potassium hydro.xide. 

 Dissolve ppt. by adding 26% ammonia 

 drop by drop and make up with abs. 

 ale. to 100 cc. Allow to stand over 

 night before use. 



Fix livers of guinea pigs and placentas 

 of same and other animals for 6-12hrs. 

 in sat. picric acid in abs. ale, 90 cc. 

 and neutral formaldehyde, 10 cc. 

 Wash in abs. ale. several times likewise 

 in chloroform and abs. ale. Transfer 

 to chloroform and embed in paraffin. 



Place sections in 0.25% aq. potassium 

 permanganate, 5-10 min.; rinse in aq. 

 dest. 1-2 min., decolorize in 5% aq. 

 oxalic acid, 5 min. and rinse again in aq. 

 dest. Place in 2% aq. silver nitrate, 

 12-24 hrs., transfer to ammoniacal sil- 

 ver nitrate, 15-30 min., rinse in 4% 

 neutral formalin, 5-20 sec. and in run- 

 ning water, 1 min. Fix in 5% aq. 

 sodium thiosulphate, 5-10 min. After 

 washing in running water, 1 min., 

 counterstain in paracarmine (Mayer), 

 dehydrate, clear in xylol and mount in 

 balsam. Glycogen, dense black corre- 

 sponds with that shown by Bauer- 

 P'eulgen technique. Excellent illus- 

 trations. 



Glycol Stearate. As an imbedding medium 

 (Cutler, O. I., Arch. Path., 1935, 20, 

 445-446). Pass up through alcohols to 

 equal parts 95% ale. and glycol stearate 

 in incubator at 56 °C. 12-24 hrs. Pure 

 glycol stearate at 56 °C. 24 hrs. Imbed 

 as in paraffin. 



Glyoxal. As substitute for formaldehyde 

 in tissue fixation (Wicks, L. F. and Sunt- 

 zeff, v.. Science, 1943, 98, 204; Stowell, 

 R. E. and Stokes, J. M. J. Tech. Meth. 

 and Bull. Internat. Assoc. Med. Mu- 

 seums, 1944, 24, 25-30). Concentra- 

 tions 2-6% produce less shrinkage and 

 give better cytoplasmic preservation 

 than 4% formaldehyde. Glyoxal is 

 only recommended as general substi- 

 tute for formaldehj'de when latter is 

 not available. 



Gmelin's test for bile pigments. On addi- 

 tion of nitric acid containing a little 

 nitrous acid, color changes to green, 

 then red and finally blue observable 

 under microscope. 



Gold, microcheraical detection of: 1. Method 



