Production of Acid by Tissues. 161 



have been no summation with the subminimum central effect of 

 the ether. 



ioo (796) 



Note on the production of acid by tissues growing in vitro. 



By Peyton Rous, M.D. 



[From the Laboratories of the Rockefeller Institute for Medical 



Research.] 



Connective-tissue cells of the chicken, growing in vitro in 

 chicken plasma to which a little blue litmus has been added, 

 produce rapidly a focal, pink coloration of the medium. If a 

 number of small fragments of one tissue (heart muscle or the aorta 

 of young chicks, or chicken sarcoma) be plated out with the 

 plasma medium in a petrie dish, it will be found that all the tissue 

 bits are at first stained blue, but that those from which growth 

 occurs become pink, while the growing tissue itself is unstained. 

 The fragments remaining permanently inert keep the blue color. 



Often a pink coloration of tissue bits can be observed at a time 

 when growth is found, microscopically, to have barely started. 

 The acid change is in general sharply localized to the neighborhood 

 of the growing tissue. When growth is checked by placing the 

 preparation in the ice-box, neutralization in the acid foci is often 

 incomplete at the end of forty-eight hours, and this even when the 

 bulk of alkaline plasma is relatively large and its plasma network 

 thinned by dilution. Diffusion in the plasma medium as thus 

 indicated is very slow. Under the ordinary circumstance of 

 in vitro life without artificial provision for a circulation of fluid, 

 tissue proliferation must take place almost from its beginning, 

 in an acid medium. This constitutes a serious fault in the method 

 of cultivation. 



The nature of the acids produced by the growing tissue has not 

 been determined. Carbonic and lactic acids are presumably 

 present in greatest quantity. That the amount of acid formed 

 may be very considerable has been shown by titrating out the 

 as yet unclotted blue plasma to the tint acquired by the tissue 

 cultures. The acid does not affect methyl orange, but very 

 occasionally it changes congo red toward violet, a change best seen 



