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Protocol for isolating and characterizing human vitreous immune cell infiltrates by flow cytometry and single-cell
Sandhya Rani Bhanja1, Sayantan Ghosh1, Pankaj Kumar2
1Ocular Immunology Laboratory, Prof Brien Holden Eye Research Centre, LV Prasad Eye Institute, Kallam Anji Reddy Campus, Hyderabad, India.
Insights
This study details a new protocol for isolating immune cells from eye vitreous. This method allows for detailed analysis of intraocular immune responses without eye damage.
Area of Science:
- Ophthalmology
- Immunology
- Molecular Biology
Background:
- Immune cells in the eye's vitreous humor offer insights into intraocular immune responses.
- Accessing these cells without harming the eye is crucial for research.
Purpose of the Study:
- To present a customized protocol for isolating and characterizing immune cell infiltrates from human vitreous samples.
- To optimize immune cell yield for downstream analyses.
Main Methods:
- Surgical modifications for enhanced vitreous sample collection.
- Flow cytometry for immune cell characterization.
- Single-cell transcriptomics for in-depth analysis.
Main Results:
- The protocol effectively isolates and characterizes vitreous immune cells.
- Maximized immune cell yield is achieved through specific measures.
- The method preserves the eye's structural and functional integrity.
Conclusions:
- The developed protocol provides a valuable tool for studying intraocular immunity.
- This method enables detailed analysis of immune cell infiltrates in various ocular conditions.
- Further research can utilize this protocol to understand immune-mediated eye diseases.
Abstract:
Vitreous infiltrating immune cells represent an accessible tissue sample that can provide valuable information on the intraocular immune response without compromising the structural and functional integrity of the eye. Here, we present our customized protocol for isolating and characterizing vitreous immune cell infiltrates by flow cytometry and single-cell transcriptomics. The protocol includes surgical modifications for sample collection and other measures to maximize the yield of immune cells from human vitreous samples. For complete details on the use and execution of this protocol, see Alam et al.1.
