NORMALITY, MICROSCOPIC 



234 



NORMALITY, MICROSCOPIC 



displaced in order to supply the neces- 

 sary illumination. But it is small with 

 relatively thick organs like the spleen 

 and liver transilluminated by Knisely's 

 quartz rod technique and examined 

 at lower magnification. With increase 

 in time modifications due to changes in 

 light, temperature, hydrogen ion con- 

 centration, etc. are likely also to in- 

 crease. 



In living cells removed from the body 

 and examined in Tissue Cultures the 

 possibility of artifact is again at a 

 minimum; but, though the cells in suc- 

 cessive generations in suitable media go 

 on living indefinitely, their environ- 

 ments are different from those existing 

 within the body. When after vital 

 staining, or supravital staining, still 

 living cells are examined in approx- 

 imately isotonic media, there is a grave 

 danger of artifact if the study is pro- 

 longed because the cells are slowly 

 dying. 



In fixed tissiies the degree of diver- 

 gence from the living condition is ob- 

 viously much greater than in the case 

 of still living ones. However death 

 has been sudden so that artifacts due 

 to gradual death are eliminated. If 

 the technique has been carefully 

 standardized the same fixative applied 

 to the same type of cell in the same 

 physiological state is likely to yield 

 similar results. 



1. The folds seen in stained sections 

 of skin are often at least partly due to 

 shrinkage of the collagenic and elastic 

 tissue of the dermis after excision and 

 before fixation. The fixative serves 

 further to exaggerate them (Evans et 

 al., 1943). Some ballooning of the epi- 

 thelial caps of intestinal villi occurs 

 in vivo. This phenomenon is likewise 

 exaggerated by ligating the arteries of 

 supply in the living animal and by em- 

 ploying fixatives which induce forcible 

 contraction of smooth muscle (Mack- 

 lins, 1932). 



2. Alterations occurring postmortem 

 in tissues before fixation can be con- 

 fusing. The most frequent one is a 

 breaking up of the tips of intestinal 

 villi, sometimes also caused by mechan- 

 ical injury during excision or washing 

 out the contents of the gut. In all 

 tissues autolytic and osmotic changes 

 occur after death which profoundly 

 alter their structure after fixation. 

 The rate of autolysis is very rapid in 

 organs like the pancreas rich in enzymes 

 let loose when the action of said en- 

 zymes ceases with death to be properly 

 regulated. The rate is slower in the 

 nervous system and especially in the 

 walls of elastic arteries in which the 



proportion of inanimate components 

 (elastic and collagenic fibers) is high 

 and that of living cells is low. 



3. Shrinkage, and increased affinity 

 of surface cells for stains, occasioned by 

 letting the surface dry before fixation 

 should be recognized as such. 



4. Differences in the appearance of 

 the peripheral and central parts of 

 tissues, owing to the unequal penetra- 

 tion of the fixative as a whole, or of its 

 components, are sometimes encount- 

 ered. 



5. The displacement of glycogen in 

 the direction of entrance and passage 

 of the fixative, so well illustrated in 

 a book by de Robertis and his associates 

 (1948), suggests the likelihood of similar 

 artifacts in the cytoplasmic localization 

 of other substances. 



6. The glassy appearance of nuclei 

 and cytoplasm, sometimes occasioned 

 by overheating in paraffin imbedding, 

 or in spreading out sections, is easily 

 detected. 



7. The presence of formed material 

 within blood vessels, faintly resembling 

 organisms, caused by the coagulation 

 of blood proteins, has led some people 

 astray. 



8. The introduction of extraneous 

 substances present either in the al- 

 bumin fixative used to mount the sec- 

 tions or deposited as dust from the air 

 (bacteria, spores, mineral matter, etc.) 

 is a possibility to be borne in mind. 

 Careful focussing is required to deter- 

 mine whether particulate matter is 

 located below, within or upon the sec- 

 tion. In stained smears of blood and 

 other cells precautions against ex- 

 traneous substances are also indicated. 



Interpretation: One has to be almost 

 constantly on guard because the fore- 

 going list of artifacts is by no means 

 complete. It is simply illustrative of 

 some of the conditions that may be met 

 with and must be excluded. Even when 

 appearances, that do not seem to be 

 artifacts, are observed it is not safe to 

 conclude in an unqualified fashion that 

 the tissue, or organ, from which the 

 preparations have been made was in 

 fact normal. On the contrary several 

 conditions require fulfillment before 

 an opinion should be offered. 



1. The appearance noted must be 

 present in more than 50 per cent of in- 

 dividual tissues comprising a homo- 

 geneous series in the sense already ex- 

 plained. The advice of a colleague 

 versed in statistics is desirable as to 

 the number required in the said series. 



2. In the event that the appearance 

 in question is observed in but small 

 pieces excised from large organs or 



