CHOICE OF JIETHODS 9 



regions of the body. Only since a technique has been devised whereby whole 

 Epidermis freed from dermis can be obtained in a condition suitable for analysis, 

 not having been exposed to any fluids, has progress been possible. 



Results of direct chemical analysis of any tissue may be misleading unless 

 interpreted in terms of its structural make up and of what has happened to it 

 since it existed in vivo. Among the experimental errors to be guarded against 

 are variability in sacrificing the animal, or the manner of death of the patient, 

 in excision of tissue allowing more or less blood and other fluids to drain out or 

 evaporate, in time and in temperature, in age, sex, and in conditions before 

 death. 



The extracellular and intracellular fluids or phases, are large in volume, when 

 all are taken together, but difficult to get at directly. To obtain data "the 

 deducive histochemical method" is suggested. This is described by Lowry, 

 0. H. and Hastings, A. B. in Cowdry's Problems of Ageing, 1942, 728-755. 



Those wishing to analyse extremely small volumes of fiuid ^^ hich by contrast 

 can be collected for direct determinations cannot do better than to familarize 

 themselves with the techniques elaborated by A. N. Richards and his associates 

 at the University of Pennsylvania for the study of glomerular urine. 



By the useful technique of Microincineration minerals which are not volatiHzed 

 at high temperature can be directly studied in the tissues in the positions which 

 they previously occupied in living organisms. They appear as shining particles 

 when viewed by the Dark Field Microscope. Microincineration is truly a 

 microchemical method for the localization of stnicturc which is microscopic 

 in its fineness. 



Quite a number of Microchemical Reactions capable of demonstrating the 

 precise location in the cells of minerals, fats, carbohydrates and proteins are 

 available. 



By a Photoelectric Microphotometer it is possible to estimate quantitatively 

 reactions like that of Feulgen for Thymonucleic Acid which give distinctive 

 colors and numerous stains which are specific for tissue components and can 

 be standardized in their action. But the data obtained are relative, that is 

 it can be said that the reaction is say 60 per cent greater in one specimen than 

 in another. The absolute amount of the component demonstrated per gram 

 of tissue cannot yet be arrived at. 



Several Enzymes (phosphatase, dopa-oxidase, arginase) can now be micro- 

 scopically identified and their position within cells determined. By close com- 

 parison of enzymatic properties with the cellular composition of tissues, the 

 localization of many others can be inferred. 



In the case of these and other microchemical ir.ethods the treatment of the 

 tissue after excision and before the special procedures are commenced is of con- 

 sequence. Even in the preparation of routine frozen sections, and much more 

 so when the specimens are fixed, dehydrated, cleared, imbedded and sectioned, 

 there are many opportunities for the loss of chemical substances and of change 

 in their position in the tissue and within cells. The best way to hold the com- 



