LYMPHATICS IN FROG LARVAE 9 



seen. The variations shown in a number of different injected 

 specimens are also very helpful in establishing a normal type. 

 At times it is difficult to secure complete injections of a 

 region, especially in early stages, but with persistence tin- 

 approximation to completion is always closer than is possible 

 by any other method, while the resulting pictures are positive 

 and objective. 



INJECTION METHOD 



The injection masses used were India ink or Berlin blue, 

 introduced with the aid of a micro-injector devised by me in 

 1908. Unfortunately it was found impossible to inject toad 

 embryos on account of their peculiar slimy and flaccid or 

 flexible consistency. In addition, the excessive black pigment 

 in the tissues of these forms spoils injection pictures, and this 

 is also true of R. sylvatica, though to a less extent. Hence 

 most of the material was taken from larvae of the other four 

 species of frog listed below (p. 10) because of their lighter 

 pigmentation. The progress of injection is greatly facilitated 

 when it can be observed against a light pigment-free back- 

 ground. Also after fixation when studied in cedar-wood oil, 

 in the forms mentioned there is the additional advantage of 

 increased contrast for the injected vessels as pigment gradu- 

 ally dissolves out with age in this oil. 



Young embryos of R. palustris and R. pipiens have in their 

 large size another character which makes injections of the 

 first lymphatics relatively easy, as compared with this problem 

 for the early stages of other species. Since young stages of 

 palustris are almost 2 mm. longer yet no further developed 

 than toad embryos of the same stages, it is evident that the 

 difficulties of injection and study in these technically trouble- 

 some stages are much reduced. In addition to the early de- 

 scription of the injection method (Knower, '08) the reader is 

 referred to the recent account by Knower, pages 51-61 in 

 McClung's ''Handbook of Microscopical Technique" (Mc- 

 Clung, '37). 



In continuing the work since the report of 1913-1914, I have 

 aimed at two main objectives : 1) to accumulate a considerable 



