80 HEWSON SWIFT 



narrow. Similar results for protozoan cells have been reported by Seschachar 

 and Flick.i^" 



The studies of Koenig and Stahlecker^'* indicate that tissues may vary 

 rather widely as to the ease of PNA extraction. Cat tissues fixed in 10% 

 formalin were used. At 23° PNA was removed in 1 hour from liver, but 

 only after 6 hours from nerve cells. At 4° PNA was not completely removed 

 from nerve tissue, even after 96 hours. Prolonged exposure to formalin 

 made the extraction very much more difficult. 



The extent of the perchloric acid treatment needed for PNA extraction 

 thus varies widely with different tissues and fixatives, and the proper con- 

 ditions need to be determined in each case. Under some conditions very 

 Uttle time may elapse between PNA removal and reduction in ultraviolet 

 absorption due to the removal of the DNA-purines. In addition, Pearse*'* 

 reports that perchloric acid also removes various protein, lipoprotein, and 

 glycoprotein fractions. 



2. Nuclease Extraction 



Enzymes are discussed here only in respect to their use in removing 

 nucleic acids from tissue sections. They have already been dealt with in 

 detail in Chapter 15. A number of different nuclease preparations have 

 been used,^'*^'^^'''^^* although those from beef pancreas have been most 

 studied. Ribonuclease has beer^ prepared by the method of Kunitz'^^ with 

 modifications suggested by McDonald.*^" Such preparations contain no 

 measurable amounts of proteolytic activity either in vitro or on 

 slides,^^'^^"'*^ and, at least in many cases, show no measurable activity on 

 DNA, as determined in vitro^'^^ or on slides with the Feulgen reaction.*^ 

 The partial removal of DNA from liver nuclei by high ribonuclease con- 

 centrations has been described''*^ '^^^ but is possibly ascribable to deoxy- 

 ribonuclease contamination. Contaminants may be present in commercial 

 samples of crystalline ribonuclease or can easily arise with bacterial con- 



"4 B. R. Sesachar and E. W. Flick, Science 110, 659 (1949). 



1" H. Koenig and E. Stahlecker, /. Natl. Cancer Inst. 12, 237 (1951). 



"^ A. G. E. Pearse, "Histochemistry, Theoretical and Applied," p. 120. Churchill, 



London, 1953. 

 1" R. J. Dubos and R. H. S. Thompson, J. Biol. Cheni. 124, 501 (1938). 

 138 D. Mazia and L. Jaeger, Proc. Natl. Acad. Sci. U.S. 25, 456 (1939). 

 i"M. Kunitz, J. Gen. Physiol. 24, 15 (1940). 

 i^»M. R. McDonald, J. Gen. Physiol. 32, 39 (1948). 

 "' B. P. Kaufmann, M. R. McDonald, H. Gay, K. Wilson, R. Wyman, and N. Okuda, 



Yearbook Carnegie Inst. 46, 136 (1948). 

 i« L. M. Gilbert, W. G. Overend, and M. Webb, Exptl. Cell Research 2, 138 (1951). 

 1" A. W. Pollister, M. Flax, M. Himes, and C. Leuchtenberger, J. Natl. Cancer Inst. 



10, 1349 (1950). 

 1" A. W. Pollister, J. Cellular Comp. Physiol. 38, Suppl 1, 87 (1951). 



