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Updated: Apr 28, 2026
3D Visualization of Retinal Vascular Pericytes in Mice by Immunostaining
Published on: November 1, 2024
Purification of pericytes from rodent optic nerve by immunopanning
Lu Zhou1, Fabien Sohet2, Richard Daneman2
1Department of Neurobiology, Stanford University School of Medicine, Stanford, California 94305-5125.
Abstract:
This protocol describes the use of immunopanning to purify rodent pericytes from the optic nerve. Immunopanning permits the prospective isolation of pericytes from optic nerve tissue by relying on the binding of pericytes to an anti-PDGFRβ (platelet-derived growth factor receptor beta) antibody adhered to a Petri dish. The cells are viable at the end of this gentle procedure, and they can be analyzed acutely for gene expression or cultured alone or in coculture with other central nervous system (CNS) cell types, including CNS endothelial cells and CNS astrocytes. As written, this procedure is used for isolation of optic nerve pericytes from the rat. The same PDGFRβ antibodies can be used for purifying optic nerve pericytes from the mouse, but alternate negative panning antibodies must be used to ensure that astrocytes do not contaminate the preparation. This procedure can also be modified to purify pericytes from the brain. The same PDGFRβ antibody is used, but additional steps (specific dissections or negative panning) are required to ensure that other PDGFRβ-positive contaminants, including cells from the rostral migratory stream, are depleted from the cell suspension.
Insights
This study details a gentle immunopanning method to isolate viable rodent pericytes from the optic nerve. This technique allows for subsequent gene expression analysis or cell culture for central nervous system research.
Area of Science:
- Neuroscience
- Cell Biology
- Immunology
Background:
- Pericytes are crucial cells in the central nervous system (CNS), playing vital roles in blood-brain barrier maintenance and neurovascular development.
- Accurate isolation of pericytes is essential for studying their function and contribution to CNS health and disease.
- Existing methods for pericyte isolation can be complex or yield cells with compromised viability.
Purpose of the Study:
- To describe a refined immunopanning protocol for the prospective isolation of rodent optic nerve pericytes.
- To ensure the viability of isolated pericytes for downstream applications such as gene expression analysis and cell culture.
- To provide a adaptable method for pericyte isolation from different CNS regions and species.
Main Methods:
- Immunopanning technique utilizing anti-PDGFRβ (platelet-derived growth factor receptor beta) antibody-coated Petri dishes.
- Prospective isolation of pericytes based on PDGFRβ expression.
- Adaptations for mouse pericyte isolation and purification from brain tissue, including strategies to avoid astrocyte and rostral migratory stream cell contamination.
Main Results:
- Successful isolation of viable pericytes from rat optic nerve tissue.
- Demonstrated adaptability of the protocol for mouse optic nerve pericyte purification with modifications.
- Outlined necessary adjustments for purifying pericytes from the brain, addressing potential contaminants.
Conclusions:
- Immunopanning offers a gentle and effective method for isolating viable rodent pericytes from the optic nerve.
- The protocol can be modified for pericyte isolation from mouse optic nerve and brain tissue.
- This method facilitates further research into pericyte biology and function within the CNS.
