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    • CommentAuthorpawhitman
    • CommentTimeJun 13th 2008 edited
     
    This protocol explains how to: Expose Xenopus Tadpole brains

    Protocol revised: 6/13/2008

    Protocol written by: Patrick Whitman



    Things needed:
    - 1x MBS
    - "Pin-on-a-stick"
    - Blunt forceps
    - Sylguard dish for pins
    - 6-8 0.2mm straight pins
    - 6-8 0.2mm bent pins

    Protocol:
    1. Tadpoles should be at stage 33/34
    2. Fill Sylguard dish with 1x MBS
    3. Put tadpoles in Sylguard dish and wait for the tadpoles to become unresponsive from the anethesia in the MBS.
    4. Pin tadpoles down using forceps and 0.2mm pins. One straight pin through the tadpole's spine. One bent pin across the body just below the head to hold it in place.
    5. Be sure to arrange the tadpoles in the dish in an organized fashion. ie, put the tadpoles in two rows, all facing the same way, same side up.
    6. Use your "pin-on-a-stick" to gently remove the layer of skin covering the eye and the brain.
    7. Peel the skin back until the entire brain is exposed, and if possible try to remove the skin you peeled back entirely so it doesn't grow back overnight.
    8. Starting at the eye, remove the pia layer covering the brain being careful not to go too deep and possibly puncture the brain.
    9. Remove the pins with your forceps and transfer the tadpoles to a 35mm petri dish filled with 1x MBS
    10. Store tadpoles in 1x MBS overnight at room temperature for HRP fill the next day.