144 Microscopic Histochemistry 



ters"^-^^ where a transient high concentration of phosphate 

 ions may prevail for relatively long periods. This type of arti- 

 fact will be accentuated on prolonged incubation. The sec- 

 ondary staining of nuclei, themselves inactive, is a typical 

 example of a false localization,^^- ^^ that is, a positive reac- 

 tion due to enzyme contained in the tissue proper but appear- 

 ing at sites other than those of the true primary microscopic 

 localization. 



In a recent paper Johansen and Linderstr0m-Lang^^ give a 

 quantitative evaluation of diffusion artifacts in the calcium- 

 cobalt technique for alkaline phosphatase. They come to the 

 conclusions that ( 1 ) calcium phosphate has a great tendency 

 to form supersaturated solutions, and (2) the time required 

 for the phosphate concentration to reach the critical level 

 may vary from approximately 0.1 to 2.5 seconds. This time 

 is ample to allow gross diffusion into the neighboring areas, 

 where conditions may be more favorable for precipitation 

 (the presence of preformed crystal nuclei; special chemical 

 or physical configuration of cell structures ) . Therefore, it is 

 very likely that calcium phosphate will precipitate at such 

 preferential sites rather than in centers of high enzymatic 

 activity. 



While the possibility of diffusion artifacts cannot be de- 

 nied, the conclusions of Johansen and Linderstr0m-Lang are 

 based on purely theoretical assumptions, some of which are 

 not borne out by experimental facts and which make the 

 situation look much worse than it actually is. For instance, 

 the authors do not take into account the results of experi- 

 ments showing that diffusion artifacts occur mainly under 



7. DanieUi, J. F.: J. Exper. Biol., 22:110, 1946. 



8. Jacoby, F., and Martin, B. F.: Nature, 163:875, 1949. 



9. Martin, B. F., and Jacoby, F.: J. Anat., 83:351, 1949. 



10. Feigin, L, Wolf, A., and Kabat, E. A.: Am. J. Path., 26:647, 1950. 



11. Kroon, D. B.: Acta endocrinol., 2:227, 1949; Doyle, W. L.r Am. J. 

 Anat., 87:79, 1950; Cleland, K. W.: Proc. Linnean Soc. New South Wales, 

 75:54, 1950; NovikofiF, A. B.: Science, 113:320, 1951. 



12. Gomori, G.: J. Lab. & Clin. Med., 37:526, 1951. 



13. Johansen, G., and Linderstr0m-Lang, K.: Acta chem. Scandinav., 

 5:965, 1951. 



