106 KANSAS UNIVERSITY SCIENCE BULLETIN. 



In making preparations of this kind, where deeply pigmented 

 specimens are used, it is often found desirable to bleach them. 

 Beautiful preparations of the segmentation stages of the frog 

 can be made by the methods explained above in which the ma- 

 terial has been prepared by immersion in hydrogen peroxide — 

 the ordinary commercial strength can be used. It is only ap- 

 plicable with specimens that have been thoroughly hardened 

 in any of the ordinary hardening fluids, or even in formalin. 

 In the latter case care must be taken with delicate structures 

 to wash out the formalin before putting them in the hydro- 

 gen peroxide, otherwise the tissues would be distorted by the 

 oxygen liberated. Frog eggs may be left in the hydrogen per- 

 oxide for even two or three weeks, if necessary. My experi- 

 ence has been that about a week will suffice for the bleaching 

 of the pigmented area to a light brown color. 



When this has been accomplished to the right stage, the 

 segmentation furrows show with remarkable clearness. One 

 great difficulty in using the frog as a type form for a study 

 of elementary embryology has been the fact that it has ap- 

 peared impossible to make whole mounts of the young tad- 

 poles. Well-fixed specimens in all stages of development can 

 be completely bleached with hydrogen peroxide and rendered 

 as white as chick embryos. They can then be stained and 

 mounted in balsam. My experience has been that a very dilute 

 stain of alum cochineal gives the best results, although I have 

 also used a dilute stain of Conklin's hematoxylin. Emphasis 

 must be laid upon the dilution of these stains. In the case of 

 both, they are diluted with distilled water until they show but 

 a faint tint of color. 



The tadpole is left in the stain over night, and is then run 

 up through the increasing grades of alcohol, being finally 

 transferred into one of the better clearing oils such as creo- 

 sote or synthetic oil of wintergreen. The tadpole is then 

 mounted in Canada balsam or dammar. 



These preparations have a very great advantage over chick 

 embryos, in the fact that no parts are missing; there is no 

 distortion due to imperfect fixation, and there is no torsion 

 of the body. If the preparation is properly carried out, they 

 should be just as clear and as easily studied as are whole 

 mounts of the chick. The brain, pituitary ingrowth, eye, 

 auditory vesicle, pronephros, heart, stomodseum and notocord 



