Obtaining Human Microglia from Adult Human Brain Tissue

Ishan Agrawal; Shivanjali Saxena; Preethika Nair; Deepak Jha; Sushmita Jha

doi:10.3791/61438

Obtaining Human Microglia from Adult Human Brain Tissue

JoVE Journal
Neuroscience
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JoVE Journal Neuroscience

Obtaining Human Microglia from Adult Human Brain Tissue

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09:41 min

August 30, 2020

DOI: 10.3791/61438-v

Ishan Agrawal¹, Shivanjali Saxena¹, Preethika Nair¹, Deepak Jha², Sushmita Jha¹

¹Department of Bioscience and Bioengineering,Indian Institute of Technology Jodhpur, ²Department of Neurosurgery,All India Institute of Medical Sciences Jodhpur

Overview

This protocol outlines a method for efficiently isolating primary microglia from live adult human brain tissue collected during surgery. The protocol emphasizes cost-effectiveness and robustness, allowing for further study of cellular processes in both homeostasis and disease contexts.

Key Study Components

Area of Science

Neuroscience
Cell Biology
Microglial Research

Background

Microglia are the resident immune cells of the central nervous system.
They play crucial roles in maintaining homeostasis and responding to injury.
Isolation of primary human microglia is essential for studying their functions.

Purpose of Study

To provide a robust method for isolating primary human microglia.
To reduce the number of steps in existing protocols to minimize cell loss.
To facilitate the use of human microglia in experimental settings.

Methods Used

The method involves cell culture techniques using live adult human brain tissues.
Tissues are collected, diced, and treated with trypsin EDTA to isolate cells.
The protocol includes critical steps like maintaining tissue temperature to prevent cell death.
Cells are cultured in a suitable medium and characterized using immunocytochemistry.

Main Results

The protocol effectively isolates primary human microglia with a high yield.
About 80% of cultured cells were identified as microglia, confirming isolation success.
Cell characterization using immunocytochemistry revealed clear distinctions between microglia and other cell types.

Conclusions

This study demonstrates an efficient method for isolating microglia, enabling further research.
The method's robustness enhances its applicability for downstream experiments in neuroscience.
Understanding the role of microglia in health and disease could improve therapeutic strategies.

Frequently Asked Questions

What are the advantages of this isolation method?

This method is cost-effective and reduces cell loss by minimizing the number of procedural steps compared to traditional protocols.

How is the brain tissue handled during the process?

Brain tissue is collected in ice cold PBS and processed immediately to prevent cell death, ensuring efficient isolation.

What types of cells are obtained from this protocol?

The protocol primarily yields microglia, with a purity of approximately 80% as confirmed by immunocytochemistry.

How long does the entire isolation procedure take?

The complete microglia isolation procedure can be accomplished in about one and a half hours.

Can this protocol be adapted for other cell types?

While the protocol is specifically designed for microglia, similar techniques may be adapted for other primary cell types with modifications.

What are the key limitations of this method?

One limitation includes the need for fresh brain tissue, which may not always be readily available.

How can the isolated microglia be used in further experiments?

Isolated microglia can be used to study their function in various experimental setups, including disease models and neuroinflammatory responses.

This protocol is an efficient, cost effective and robust method of isolating primary microglia from live, adult, human brain tissue. Isolated primary human microglia can serve as a tool for studying cellular processes in homeostasis and disease.

The overall goal of the protocol is to isolate primary microglia cells from live adult human brain tissues. In our case, this was collected as surgical window during surgery. This is accomplished by initially collecting the brain tissues in ice cold PBS, the tissue is then diced into 1-mm cube small pieces, and I just did with the help of trypsin EDTA.

Following this, the cells are harvested by centrifugation and plated in a flask suitable for androgen cell culture for 48 hours. Cells are harvested once more from the flask and cultured till further use. Primary human microglia cells can now be characterized by using immunocytochemistry and used for further experiment.

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