86 HEWSON SWIFT 



known must be involved. A shift to 290 mn is also caused by certain chemi- 

 cal combinations of the phenolic groups, for example, with mercuric ions. 

 Oxidation of the phenolic group causes marked changes in the shape of 

 protein absorption curves, although the position of the peak is un- 

 changed.^*^ Some of these effects on nucleic acid or protein absorption prob- 

 ably do not influence cytochemical determinations; others, however, such 

 as the effect of depolymerization or tyrosine oxidation, may be produced by 

 fixation or embedding of material, and their influence needs investigation, 

 particularly if accurate nucleic acid determinations are desired. 



2. Interfering Substances 



In addition to protein absorption, there are a number of other cell con- 

 stituents that absorb in the 260-m/Lt region, which may interfere with 

 nucleic acid determination. Nucleotides such as ATP or DPN absorp 

 ultraviolet light, but it is doubtful if these occur in sufficient quantity to 

 have noticeable effects on absorption curves of most cells. During the 

 process of fixation and embedding, such compounds are probably readily 

 extracted. In plant cells both catechol and ascorbic acid may add to ab- 

 sorption in the nucleic acid region. Catechol is readily removed by fixation, 

 but ascorbic acid was found to remain in bean root cells after acetic alcohol 

 or absolute alcohol fixation. i^" 



A yellow pigment, possibly a pterin, with strongly absorbing 260- and 

 280-m/i maxima occurs in human nervous tissue. The pigment is not readily 

 extracted and so remains in tissues after cytological preparation. ^^^ Such 

 substances are probably of little consequence in the great majority of 

 tissues, but should obviously be watched for in any new material. Their 

 presence is usually recognizable where whole absorption curves are run 

 through the region of specific nucleic acid absorption (230 to 320 m^i), and 

 where nuclease-treated blanks are also measured. 



3. XoNSPEciFic Light Loss 



Nonspecific light loss is particularly severe in the ultraviolet region. 

 Light scattering is produced by the many changes of refractive index in the 

 fine structure of the tissue, and its magnitude increases with smaller wave- 

 lengths at a rate dependept upon the dimensions and refractive index of 

 the scattering particles. Since the submicroscopic structure that constitutes 

 living or fixed protoplasm is obviously heterogenous and the particle dimen- 

 sions are not readily measured, the slope of the scatter curve cannot be 

 9xperLmentally determined. The extent of scatter at 320 to 350 m/x can be 



89 E. S. G. Barron and P. Finkelstein, Arch. Biochem. and Biophys. 41, 212 (1952). 

 «J. Chayen, Symposia. Soc. Exptl. Biol. 6, 290 (1952). 

 H. Hyden and B. Lindstrom, Discussions Faraday Soc. 9, 436 (1950). 



