10 Development and fciliajje oi' I'riniferous Tubules 



Method. 



Of the various fixinc: fluids used, fixation by Zenker's fluid proved most 

 satisfactory. 



The wax-plate reconstruction method \yas largely used in gaining the 

 data which form the basis for this contribution. It was found that the 

 material could be most readily manipulated in this way by using serial 

 sections 5 /x. in thickness and a magnification of 400 diameters, necessitat- 

 ing the use of wax-plates 2 mm. in thickness. As already stated, it is 

 often exceedingly difficult in sections more than 5 /a in thickness to 

 interpret with any degree of clearness the earlier stages in the develop- 

 ment of the kidney tubules ; the same may be said of sections from more 

 fully developed kidneys, where the tubules are already well differentiated. 

 The difficulty here met with is that even in sections having a thickness 

 of not more than 10 /x, the tracing of a tubule through a series of sections 

 is often made difficult by the fact that portions of two tubules lying over 

 each other— with reference to the plane of the section — and in contact 

 are both included in the same section and appear as one tubule, thus 

 easily leading the observer astray. For the earlier stages — embryos 

 having a length of about 1.5 cm. — the entire embryo was cut in sagittal 

 serial sections; for embryos measuring from 1.5 to 2.5 cm., only the 

 posterior half was thus cut, and for older stages, the kidneys were 

 removed after fixation and cut in series either longitudinally or trans- 

 versely. The difficulty of obtaining unbroken series of sections 5 fi in 

 thickness, without wrinkling or distortion when mounted, was met for 

 many of the series by cutting the sections one by one on an ordinary 

 sliding microtome — Altmann pattern — with the knife at an angle of about 

 45° and with the knife blade covered with a layer of distilled water while 

 cutting. This latter procedure I have found very useful in cutting thin 

 paraffin sections. The sections are transferred as cut to an Esmarch dish 

 filled with distilled water in which they float and flatten out. They are 

 arranged as cut on a slide covered with albumin fixative placed at an 

 angle in the same dish. When the slide has been filled with sections, it 

 is placed on the warm oven until the water evaporates. The sections are 

 then ready for staining. This procedure, though much more time- 

 consuming than when the sections are cut with an automatic microtome, 

 seemed to present advantages which compensated for the additional time 

 necessary for its prosecution ; especially was this true for the older stages, 

 in which the kidneys alone were cut. The sections were stained on the 

 slide in Ehrlich's hgematoxylin and counterstained mainly in eosin or 

 erythrosin, the latter giving the better differentiation and sharper con- 

 trast than the other protoplasmic stains used. 



