Make staining solution by mixing 1 grm. carmine, O.5 grm. aluminum chloride, 2 cc. water and 100 cc. of 50 per cent alcohol, heating over the flame for 2-3 minutes until mixture darkens. Let stand 24 hours and filter. The stock solution may be diluted 1-10. Stain 10 minutes. If it does not stain well add 0.5-1 grm. of aluminum chloride. Mucin alone should be stained red. Counterstain with hæmatoxylin.
XIX. MYELIN. This appears in the form of doubly refractive granules, that stain with less intensity with the fat dyes, but may be differentiated from fat in that it loses the power of reducing osmic acid after being mordanted for eight days or more in bichromate solutions, while fat does not.
XX. NECROSIS. Hæmatoxylin and eosin, and Van Gieson’s give good pictures. Use Weigert’s fibrin stain for coagulation-necrosis, and Benda’s method for the demonstration of fatty acids for the staining of fat-necrosis. Recent necrotic areas stain diffusely blue with hæmatoxylin; older areas may take the plasma stains alone. Use various methods for the demonstration of micro-organisms in the necrotic areas.
XXI. NEOPLASMS. Use hæmatoxylin and eosin, and Van Gieson’s for ordinary diagnosis. To differentiate sarcoma and carcinoma use Van Gieson’s, Mallory’s or other reticulum stains. For the study of cell-inclusions use Altmann’s, Russell’s, Plimmer’s and Pianese’s methods. Special fixation or Zenker’s is necessary. Methylene-blue and eosin after Zenker’s give excellent pictures. For the demonstration of mitoses the methods given above should be employed.
XXII. PIGMENT. Use the carmines for contrasting melanin, hæmofuscin, lipochromes, hæmatoidin, hæmosiderin, bilirubin and all yellow, brown, blue, black, etc., extrinsic pigments. In tissue fixed in mercuric chloride or formol bilirubin is green, and can thus be differentiated from hæmatoidin. The lipochromes give weak fat-reactions, and this is used to distinguish them from other yellow or brown pigments. Alcohol fixation is the best for pigment study, although the other fixing solutions may be used. Formol sometimes produces pseudo-pigments by its action upon hæmoglobin. The iron-reactions are obtained best in sections cut on the freezing-microtome, although both paraffin and celloidin imbedding may be used. In testing for iron glass needles should be used and all traces of iron should be removed from staining-dishes, slides, etc., by treating with hydrochloric acid, distilled water and alcohol.
1. Potassium Ferrocyanid Test for Iron.
1. Stain sections in lithium carmine for several hours.
2. Differentiate in acid alcohol, stopping short of the desired complete differentiation of the nuclei.
3. Wash in water.
4. Saturated solution of potassium ferrocyanid 1-3 hours.