Cerebellum Dissection and Slice Preparation: A Method to Generate Tissue Slices from Mouse Cerebellum

Published: April 30, 2023

Abstract

Source: Thetiot, M. et al. Preparation and Immunostaining of Myelinating Organotypic Cerebellar Slice Cultures. J. Vis. Exp. (2019)

This video describes a method to generate tissue slices from the mouse cerebellum. The cerebellar slices can be used for organotypic culturing that provides insights into neurodevelopmental mechanisms, including myelination dynamics.

Protocol

All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board. 1. Cerebellum Dissection and Sections Preparation (Hands-on Time ≈ 15–20 min per animal) Place one of the 60 mm cell culture dishes containing the ice-cold dissection medium under the binocular microscope and remove the top plate. Swab the head fur with 70% ethanol (carefully avoiding …

Representative Results

Figure 1: Illustration of cerebellum slices generation. The dissection is divided in three main steps. (A) The cerebellum is isolated, and the meninges removed (steps 1.10 and 1.11). (B) The cerebellum is then transferred to the chopper platform and sliced (steps 1.12 to 1.16). (C) Lastly, the slices are isolated from the vermis …

Açıklamalar

The authors have nothing to disclose.

Materials

BME medium ThermoFisher Scientific 41010026
Hank’s Balanced Salt Solution (10x HBSS) ThermoFisher Scientific 14180046
GlutaMAX (100x) ThermoFisher Scientific 35050038
Heat-inactivated Horse Serum ThermoFisher Scientific 26050088
Penicillin–Streptomycin (10.000 IU/mL) ThermoFisher Scientific 15140122
Gey’s Balanced Salt Solution Sigma Aldrich G9779-500ML
D-Glucose Solution (45%) Sigma Aldrich G8769-100ML
Paraformaldehyde (PFA) Electron Microscopy Sciences 15714
Absolute ethanol (100% ethanol) VWR Chemicals 20821.33
Triton® X-100 Sigma Aldrich X100-500ML Cooled at -20°C
10% Normal Goat Serum (NGS) ThermoFisher Scientific 500622
Phosphate Buffer Solution EuroMEDEX ET330-A
Tissue chopper McIlwain pH 7.4
Razor blades
Large scissors F.S.T 14101-14
Small scissors F.S.T 91500-09
Fine-straight forceps F.S.T 91150-20
Curved-fine forceps F.S.T 11297-00
Cell culture dishes (60 mm and 100 mm) TPP
4-, 6-well culture plates TPP
Millicell culture inserts (0,4 µm, 30 mm diameter) Merck Millipore PICM0RG50
Cell culture incubator
Fine-end pipette tips Dutscher 134000CL 37°C, 5% CO2
Wide-bore pipette tips ThermoFisher Scientific 2079G
Sterile syringe Terumo Europe
Sterile syringe filters Terumo Europe 20 or 50 mL
Scalpel Swann-Morton 510 0.22 µm
Brush
Microscope slides RS France
Glass coverslips RS France 76 mm x 26 mm x 1.1 mm
Kimtech Sciences Tissue Wipers Kimberly-Clark Professional 5511 22 mm x 22 mm
Binocular microscope

Etiketler

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Bu Makaleden Alıntı Yapın
Cerebellum Dissection and Slice Preparation: A Method to Generate Tissue Slices from Mouse Cerebellum. J. Vis. Exp. (Pending Publication), e20696, doi: (2023).

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