470 Prof. A. B. Macallum. Detection and Localisation 



The error at the base of the process adopted by Lilienfeld and 

 Monti has been the assumption that it is possible to wash out of 

 tissues all traces of the ammonium molybdate not combined with 

 phosphoric acid. I have found that when the stamens of Erythro- 

 nitim americanum, treated for twenty-four hours with the nitric- 

 molybdate solution, were washed with distilled water many times for 

 five months, they gave, at the end of that time, marked evidence of 

 the presence in them of ammonium molybdate. The addition of 

 stannous chloride brought out everywhere in such preparations the 

 appearance of the blue molybdic oxide, whereas, when such prepara- 

 tions were treated with pyrogallol solution, the phospho-molybdate 

 compound was found to be limited in its distribution. From animal 

 tissues also, I have found it impossible to remove by washing the 

 molybdate reagent. Indeed, one may succeed thus in removing the 

 phospho-molybdate compound rather than the other. 



Heine* also has specially insisted on the strong affinity that cell 

 substances, those which contain phosphorus as well as those which 

 do not, have for ammonium molybdate, forming with the latter, 

 compounds insoluble in water or in dilute nitric acid solutions. He 

 prepared a quantity of histon, free from phosphorus compounds, 

 which, after treatment with the nitric-molybdate reagent and after 

 frequent washings, gave with stannous chloride abundant evidence of 

 the presence of ammonium molybdate. 



One is consequently justified in concluding that the results of 

 Lilienfeld and Monti's observations, based as they are on the 

 *' yellow, brown, or black " reaction of the pyrogallol, are incorrect, 

 and that while the reaction may appear in phosphorus-holding 

 elements, it is simply a coincidence, and not an indication of the 

 presence of phosphorus. 



The property of pyrogallol to form, in the reduction of the 

 molybdate and the phospho-molybdate compounds, a coloured sub- 

 stance which can be taken by cellular elements just in the same way 

 and to about the same extent that they take up other colouring 

 matters in solution, constitutes an objection to the use of this 

 reducing reagent. It is not possible to be certain in all cases in regard 

 to the length of time during which it is to be allowed to act, and, 

 consequently, a very faint green may be obscured by a light brown 

 reaction, resulting either from the oxidised pyrogallol or from the 

 reduced molybdate in the presence of traces of nitric acid. 



In consequence of this objection, I endeavoured to find a reducing 

 reagent which would leave the molybdate compound, in the presence 

 of nitric acid, unaffected, while it would markedly react with the 

 phospho-molybdate, not only in the test-tube, but in tissues. Zinc 



* "Ueber die Molybdansaure als mikroskopisches Eeagens," ' Zeit. fur Physiol. 

 Chetnie,' vol. 22, p. 132, 3896-97. 



