Kubke Lab:/Notebook/Cranial nerve development/2010/12/03

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9:45am Cut embryo RC002 with a razor blade and mounted on polylysine-subbed slides -CUT OFF HINDBRAIN MUST LEARN ANATOMY

(Note: What embryo number?--MF Kubke 04:32, 20 January 2011 (EST))
  • A mould was made by placing a small amount of OCT into the plastic mould and then the embryo was placed onto the OCT and the mould filled with OCT.
  • A mark was made on the plastic so that the orientation of the embryo could be assessed.
  • The mould was placed in the cryostat at CT-24, OT-24.
  • A chuck was placed in the cryostat and allowed to cool down for a minute before a small amount of OCT was added to the chuck.
  • As the OCT was beginning to freeze the mould was placed on top of the chuck using the marking on the plastic to ensure the correct orientation.

I then had a personal meeting with Fabiana to discuss personal objectives and progress. Emailed Satya regarding publishing her notes to OpenWetWare. -NO RESPONSE

1:40pm Began trimming the mould at 14um. 1:50pm During trimming the OCT is curling heavily

  • Changed the angle of the platform (2.5,7.5,10 degrees) - NO CHANGE
  • Added more supporting OCT to the mould. -NO CHANGE
  • Attempted to sculpt an 'anti-curling' shape in the OCT, candy-wrapper shape - NO CHANGE
  • Tried using excessive OCT - NO CHANGE
  • Tried using a razor blade to prevent the sections from curling. As the section comes off the blade I used a cold razor blade to capture the section and mount it to the slide by removing the blade from the section and quickly before it curled mounted it.

WORKED! The razor blade CAN be used to keep the sections from curling following cutting however we will need to stain them to see if the method is damaging the tissue. --Reuben Cutfield 18:42, 7 December 2010 (EST)


10.00 AM - 12.00

  • Meeting with Fabiana with regards to personal objectives.

12.00 AM- 1.10 PM

  • An incision was made at the level of the wing buds on MH002. The rostral section was placed into a mould and put into the cryostat to incubate at CT-24 OT-24.
  • Today was Reubens day to use the cryostat for that reason no cutting was done--Malisha Hettiarachchi 18:41, 12 December 2010 (EST).
  • The embryo was moved from the cryostat and into the freezer following direction from Satya. At this stage the effect of freezing an embryo once placed in the mould was not noted.

(Note: What happened to the embryo if you didn't cut it?--MF Kubke 04:34, 20 January 2011 (EST)) - I have added a note for this.