Home › Neuroscience › Vibratome Sectioning Mouse Retina to Prepare Photoreceptor Cultures
Neuroscience JoVE (Open Access) Citable · DOI

Vibratome Sectioning Mouse Retina to Prepare Photoreceptor Cultures

DOI: 10.3791/51954-v
What you'll learn
  • ✓ Isolate photoreceptor layer from mouse retina using vibratome sectioning
  • ✓ Dissociate and culture primary photoreceptor cells for analysis
  • ✓ Prepare retinal tissue for molecular, biochemical, and transplantation studies
Protocol

Neural retina of a mouse aged 8 days is on top of a 4% gelatin block. After isolation of the photoreceptor layer (200 µm) by vibratome, the photoreceptors are seeded after mechanical and enzymatic dissociation for culture. The photoreceptor layer can be used for molecular, biochemical analyses or transplantation.

Difficulty
advanced
Total time
~2–3 hours per mouse
Model organism
Mouse (postnatal day 8)
Biosafety
BSL-1

Steps

1
Prepare equipment and materials for retina sectioning

Assemble vibratome, prepare 4% gelatin embedding block, and organize dissection tools and media. Ensure proper setup before tissue isolation.

▶ 01:40
2
Isolate and flatten mouse retina from eyecup

Extract retinal tissue from enucleated eye and mount on gelatin block with photoreceptor side up. Flatten tissue gently to ensure uniform sectioning.

▶ 02:46
3
Section retina using vibratome to isolate photoreceptor layer

Use vibratome to cut approximately 200 µm thickness sections containing the photoreceptor layer. Collect sections in appropriate culture medium.

▶ 04:17
4
Dissociate and seed photoreceptor cells into culture

Treat isolated photoreceptor layer with enzymatic and mechanical dissociation to obtain single cells. Seed dissociated photoreceptors at appropriate density for culture.

▶ 06:35
5
Characterize cultured photoreceptors by microscopy

Assess photoreceptor viability, morphology, and identity using brightfield and fluorescence microscopy. Verify culture quality before downstream applications.

▶ 09:03
💬 Comments coming soon