TRANSPLANTATIONS 



PURPOSE: To determine the ability of various organ anlagen to adjust to and differentiate 

 within a new environment. 



MATEBIALS : 



Biological : Early embryos of the various amphibia, Anura and Urodela. 



Technical : Standard equipment. 



METHOD: 



Precautions : 



1. Eemove and transplant only the area and the cells prescribed. This is very im- 

 portant because the areas may vary with the age of the donor, and the various germ 

 layers may have different developmental relations to the organ, 



2. Adequate excavation of the host site must be made, especially since there is very 

 rapid healing of any embryonic wound. 



5. The transplant must "take" (become firmly attached) before the host is moved to 

 a new environment, or changed to a different medium. 



k. Twitty (1937) and others, have found that the tissues of certain amphibia produce 

 toxins which paralyze or kill the host (or transplant) in xenoplastic combina- 

 tions. Triturus tissues are particxilarly potent when combined with Amblystoma. 

 Therefore, In making xenoplastic transplants one must keep in mind the possibility 

 of tissue in compatibility. 



Controls: Since most of the organ anlagen that will be used are bilateral, the organ 

 of the unoperated side of the host may be considered as the control organ. 



Procedure : The following directions will be specific for each of the various organs. 

 It is recommended that the student consult the Chapter in this Manual pertaining to 

 the organ under study. 



Where natural pigmentations can be used to identify and trace a transplant 

 (graft) in the host environment one need not add any further marking. In homoplas- 

 tic transplants it will be necessary to pre-stain the donor tissue (graft) in Nile 

 Blue Sulphate or Neutral Bed, in order to identify and follow the fate of the graft. 

 Operated embryos should be allowed to recover In #2 Stender dishes with agar bases. 



LIMBS 



The f orelimbs of Amblystoma punctatum ( texanum, Jef fersonianum, opacum and 

 Triturus torosus) are smaller and slower growing than those of A. tigrinum and 

 A. mexicanum (axolotl). However, the A. punctatum anlage appear early (Stage #57) 

 and develop digits shortly thereafter (Stage #1+1), while the forelimb buds of the 

 A. tigrinum do not appear until about the beginning of the larval period (when the 

 yolk is resorbed). Detwiler (1958) states that the prospective limb material is 

 determined as early as the late yolk-plug stage, and Swett has shown that the two 

 axes of the limbs are laid down consecutively. (See exercise on Limb Fields.) 



i. Eemove the vitelline membranes of 10 specimens of Amblystoma (Stage #28) and 

 place them in sterile 10^ Standard Solution. If necessary, narcotize the embryos in 



1/5000 MS 222. If the transplants are to be within the same species, pre-stain the 

 donors in Nile Blue Sulphate. 



2. Select a pair of embryos of similar developmental stage and place them side- 

 by-side in an operating dish over soft paraffin or Permoplast. Mould a depression 

 for the host, and place it in the depression with the right side uppermost. 



5. Locate the forelimb anlage. This will be found ventral to somites #5-#5, 

 Just posterior to the gill swelling and includes a portion of the ventral slope of 

 the pronephric bulge. 



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