CHAPTER V 

 QUANTITATION IN HISTOCHEMISTRY 



The greatest advantage of biochemical over histochemical 

 methods is the far superior abihty of the foiTner to quantitate 

 results. However, as v^ill be show^n, a modest degree of quan- 

 titation can be achieved also by purely histochemical tech- 

 niques. 



Absorption colorimentry is the main tool of quantitation in 

 analytical chemistry, biochemistry, and histochemistry. How^- 

 ever, conditions prevailing in microscopic sections only 

 rarely permit the theoretically correct application of color- 

 imetry. 



One of the fundamental principles of absorption color- 

 imetry is that the distribution of the absorbing material in 

 the sample must be uniform. If this condition is fulfilled, 

 quantitative evaluation of photometric readings obtained 

 through the microscope is an entirely correct procedure, and 

 it has given much valuable information especially in the 

 hands of Caspersson^ and his school and a fev^ others.^ It 

 should be remarked, how^ever, that areas sufiiciently uni- 

 form in optical density are, as a rule, very small, usually 

 occupying only a minute fraction of an oil-immersion field. 

 The minimum diameter required for a measurement is about 

 four times that of the w^ave length of the light used. 



Irregular distribution of the absorbing material vv^ill lead 



1. Caspersson, T.: Arch. f. Physiol., Vol. 73, Suppl. 8, 1936, and J. Roy. 

 Micr. Soc, 60:8, 1940. 



2. Gersh, I., and Baker, R. F.: J. Cell. & Comp. Physiol., 21:213, 1943; 

 Stowell, R. E.: J. Nat. Cancer Inst, 3:11, 1942, and Anat. Rec, 91:301, 

 1945; Ris, H., and Mirsky, A. E.: J. Gen. Physiol., 33:125, 1949; Swift, 

 H. H.: Physiol. ZooL, 23:169, 1950, and Proc. Nat. Acad. Sc, 36:643, 1950; 

 and Hoover, C. R., and Thomas, L. E.: J. Nat. Cancer Inst., 10:1375, 1950. 



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