THE EFFECT OF THICKNESS OF THE SECTION 43 



to use sections which are thicker or thinner than this. When 

 this is done, it may be necessary to consider whether any of the 

 times of the various treatments should be modified. Obviously 

 a time of washing which may be satisfactory for a 2 ^ section 

 may not be satisfactory for a 16 /* section. Whether this is so 

 depends on whether the major resistance to diffusion is encoun- 

 tered in diffusing out of or into the tissue section, or whether the 

 dead layer of water surrounding the section constitutes the 

 major barrier. If the resistance to diffusion is a function of the 

 thickness of the section, then the time required to secure a given 

 degree of washing should be roughly proportional to the square 

 root of the thickness, whereas, if the dead layer of the water 

 is the main factor, the time for obtaining this given degree of 

 washing should be independent of the thickness. To investi- 

 gate this point a number of series of sections were cut at 1 /*, 2 /*, 

 4 fi, S [x, and 16 /*. These were incubated in the glycerophos- 

 phate incubation mixture for 15 minutes, and then developed 

 with the usual cobalt technique, with the exception that different 

 sections of the same thickness were washed for various lengths 

 of time in distilled water. The times for washing varied be- 

 tween 0.25 minute and 16 minutes. Owing to the solubility of 

 cobalt phosphate in distilled water, washing for more than 2 

 minutes with an 8 /x section causes loss of apparent enzyme ac- 

 tivity from a section if carried out in distilled water. The losses 

 are much smaller if the washing is carried out in 0.002 percent 

 sodium veronal, as was the case with the results displayed in 

 Table IV. Under such conditions the minimum washing time 

 for removal of cobalt nitrate increases from about 1 minute for 

 1 fx sections to about 4 minutes for 16 p sections; with 1 /* 

 sections a significant amount of cobalt phosphate was not re- 

 moved from the section even after 16 minutes' washing. 



It may perhaps be mentioned here that, whereas most tissues 

 do not take up cobalt to a significant degree from cobalt nitrate 

 solution, this is not the case when ova are studied. The yolk 

 of echinoderm ova, for example, takes up cobalt very readily: 

 this cobalt cannot be removed by washing in distilled water 

 without also removing any cobalt phosphate which is present. 

 So far no satisfactory solution has been found to this problem, 

 but it seems possible that, if a lead or copper solution is sub- 



