Kubke Lab:Research/CND/Records2010-2011Summer/MH003

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Cranial Nerve Development Experiment

Embryo details


Species:Gallus gallus domesticus
Embryo Name:MH003
Embryo stage:20-22
Staging description:Staged by Fabiana unsure of what the criteria was.
Fixation:PF
Cryoprotection:NO
Material label and storage: Stored in a vial labelled MH003 filled with PF at room temperature

Experiment details


Objective:
Procedure: Cryostat Sectioning Cryostat settings (for a more detailed protocol visit Kubke_Lab:Cryomicrotomy)

Cryostat Histology Lab
Knife
Day Cut
Knife Angle
Chamber Temp
Object Temp
Glass Slides Polysine
Plane of section Coronal
Number of slides 9
Observations Waiting for approximately 30 seconds or more after cutting a section reduced the curling of OCT and shaping the mould appropriately

(Include in your observations, eg ,were the sections serial, was any section lost, was quality assessed, etc)




Comments: Slides 1-4 cut by Fabiana, slides 5-9 cut by Malisha


The description of the staining procedure can be found here

Date 16/12/2010
Defatting and rehydration step
Solution Time Comments
Water Omitted
70% alcohol 3 min
75% alcohol Omitted
95% alcohol 3 min
100% alcohol 1 3 min
100% alcohol 2 Omitted
Xylene 1 15 min
Xylene 2 Omitted
Xylene 3 Omitted
Xylene 2 Omitted
Xylene 1 Omitted
100% alcohol 2 Omitted
100% alcohol 1 2 min
95% alcohol 2 min
75% alcohol Omitted
70% alcohol 2 min
Water 4 dips
Staining and differentiation step
Solution Time Comments
Water 4 dips
Cresyl Violet 5 min
50% alcohol Omitted
70% alcohol 2 min
70% alcohol acetic acid Omitted
95% alcohol 2
100% alcohol 1 2
100% alcohol 2 Omitted
Xylene 1 2-3 min
Xylene 2 Omitted
Xylene 3 Omitted
Coverslip

Results

Slides 1-4: Feedback from Fabiana after looking at these slides was to cut below a temperature of -27C and at 23 microns. Please refer to following entry

Images

Summary