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Related Experiment Video

Updated: Jul 7, 2025

Isolation of Adult Human Astrocyte Populations from Fresh-Frozen Cortex Using Fluorescence-Activated Nuclei Sorting

Isolation of Adult Human Astrocyte Populations from Fresh-Frozen Cortex Using Fluorescence-Activated Nuclei Sorting

Published on: April 16, 2021

Primary Astrocytes Purification and Immortalization.

Lei Qin¹, Guozhi Xiao²

¹Department of Orthopedics, Huazhong University of Science and Technology Union Shenzhen Hospital, Shenzhen, China.

Current Protocols

|December 22, 2023

Summary

This summary is machine-generated.

This study presents efficient protocols for purifying and immortalizing primary astrocytes from mouse brains. These methods yield reliable cell models for studying astrocyte biology and function in vitro.

Keywords:

SV40 large T antigen astrocytes calcium transient immunopanning primary cell immortalization

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Related Experiment Videos

Last Updated: Jul 7, 2025

Isolation of Adult Human Astrocyte Populations from Fresh-Frozen Cortex Using Fluorescence-Activated Nuclei Sorting

Isolation of Adult Human Astrocyte Populations from Fresh-Frozen Cortex Using Fluorescence-Activated Nuclei Sorting

Published on: April 16, 2021

Rapid and Refined CD11b Magnetic Isolation of Primary Microglia with Enhanced Purity and Versatility

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Published on: April 13, 2017

Isolation and Direct Neuronal Reprogramming of Mouse Astrocytes

Isolation and Direct Neuronal Reprogramming of Mouse Astrocytes

Published on: July 7, 2022

Area of Science:

Neuroscience
Cell Biology
Biotechnology

Background:

Astrocytes are crucial for central nervous system (CNS) homeostasis and function.
Primary astrocyte cultures are valuable in vitro models for studying astrocyte biology.
Existing methods for primary astrocyte isolation and immortalization can be time-consuming or costly.

Purpose of the Study:

To develop a rapid, cost-effective protocol for purifying primary astrocytes from mouse cortex and spinal cord.
To establish a method for immortalizing these purified primary astrocytes using lentivirus-mediated SV40 large T antigen expression.
To validate the utility of the immortalized astrocytes for studying astrocyte-specific markers and functions, such as calcium signaling.

Main Methods:

Modified immunopanning technique for primary astrocyte purification.
Lentivirus infection with SV40 large T antigens for astrocyte immortalization.
Assessment of astrocyte-specific marker expression.
Measurement of ATP-induced calcium flux in immortalized astrocytes.

Main Results:

Successful purification of primary astrocytes from mouse CNS tissue.
Generation of immortalized astrocyte cell lines that retain characteristics of primary astrocytes.
Demonstrated ability to measure functional responses, including calcium transients, in immortalized astrocytes.
Protocols ensure that immortalized astrocytes accurately mimic primary astrocyte cell biology in culture.

Conclusions:

The presented protocols offer efficient methods for primary astrocyte purification and immortalization.
The resulting immortalized astrocytes serve as robust models for in vitro studies of astrocyte biology and function.
These methods may be applicable to the immortalization of other primary cell types.