Kubke Lab:Research/CND/Records/MH008

=Embryo details=

Species: Gallus gallus domesticus Embryo Name: MH008 Embryo stage: ST19 Confirmed by Fabiana (supervisor). (see journal entry) Staging description: Staged by Fabiana. Fixation: Cryoprotection: Material label and storage:

=Experiment details=

Objective:To complete a serial coronal sectioning of the embryo rostral to the wingbud prior to staining with Cresyl Violet and coverslipping for histological and cytological analysis of the stained sections. Procedure: 26th Jan 2011 see Notebook entry.
 * The embryo MH008 was staged according to the and Hamilton (1951) staging system.


 * 11:30am Using dissection scissors and under a dissection microscope I cut the embryo just rostral to the wing bud
 * The head region of the embryo was then gently replaced into a vial filled with PFA using forceps.
 * A small plastic mould was half filled with OCT and then, using a blade, the embryo was very carefully laid on top of the OCT from a Petri dish containing PFA.
 * Using forceps the embryo was oriented such that the hindbrain ran parallel to the sides of the mould so that coronal sections could be made
 * Bubbles in the OCT were removed with the forceps.
 * 12:05pm Incubated the block at -19°C in the cryostat chamber.
 * 12:30pm The block was oriented 90° to a chuck and stuck on by freezing OCT between the two contacting surfaces.
 * OCT was slowly built up either side of the block inside the cryostat chamber.
 * The chuck was inserted into the metallic chuck holder and incubated at -19°C for 30 minutes.
 * 1:30pm Began trimming the block
 * Serial sections of the specimen were cut and mounted onto microscope slides and dried overnight in a staining rack, in a fume hood, wrapped in tin foil. See the Cryomicrotomy page for information on how this was done.
 * The sections were then stained using Cresyl Violet (see below for the time the slides spent in each solution). See Notebook entry. Only the staining and differentiation step was used.

Cresyl Violet staining For more informattion see Kubke_Lab:Nissl_Stain_Protocol

Comments: Staining is lighter than usual. Appears to show more contrast.

=Results= =Images= =Summary=