E, Field Operations 

 36. LIMB FIELD OPERATIONS 



PURPOSE: By the application of experimental methods to the linnb field (anlage) of the 

 Urodele embryo, to determine the 



1. Powers of regeneration following partial and total extirpation of the anlage. 



2. The effect of rotating the limb field axis. 



3. The effect of splitting the limb field. 



4. The relative functions of the ectoderm and the mesoderm in limb formation. 



5. The differentiating capacity of heterotopic transplants. 



6. The relation of host and donor in xenoplastic transplants. 



7. The genetic factors relating to limb formation, by means of 



a. Heterogenic transplants (i. e. , between species with different rates of 

 development. 



b. Heteroplastic transplants involving pigmentary differences. 



MATERIALS: 



Biological: Urodele larvae of stages #29 to #35. 



Slow developing forms Rapid developing forms 



Amblystoma punctatum (maculatum) Amblystoma tigrinum 



Amblystoma texanum (microstomum) Amblystoma mexicanum (axolotl) 



Amblystoma jeffer sonianum 



Amblystoma opacum 



Triturus rivularis 



Triturus simulans 



Triturus torosus 



Technical : Standard Operating Equipment. 



Vital dyes and MS 222 freshly made up cone. 1/3,000. 



METHOD: 



Precautions : 



1. Use 0. 1% sodium sulfadiazine if bacterial infection is encountered. 



2. Be reluctant to examine the transplant. It should remain undisturbed for 30 

 minutes or more and any necessary transferring should be by wide-mouthed 

 pipette. 



3. Temperatures lower than that of the laboratory are best for Urodele recovery 

 and survival. Some species do best at 10 C. 



4. Operations are perforn:ied in Operating Medium; embryos are later trans- 

 ferred and raised in Growing Medium. Feeding should begin when the bal- 

 ancers disappear. 



5. Muscular activity begins at about Stage #34. If operations are attempted at 

 this stage it will be necessary to use freshly made MS 222 in operating 

 medium 1/3, 000. 



Control : The controls for each of the following experimental procedures will, of 

 necessity, be different. 



Procedure : Each of the following exercises deals with the limb field but since each 

 illustrates a different principle, the procedures will naturally differ somewhat. 



* The author acknowledges, with appreciation, the help of the late Dr. F. H. Swett in organizing this exercise. 



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