title: Novel Applications of Magnetic Cell Sorting to Analyze Cell-Type Specific Gene and Protein Expression in the Central Nervous System.
abstract: The isolation and study of cell-specific populations in the central nervous system (CNS) has gained significant interest in the neuroscience community. The ability to examine cell-specific gene and protein expression patterns in healthy and pathological tissue is critical for our understanding of CNS function. Several techniques currently exist to isolate cell-specific populations, each having their own inherent advantages and shortcomings. Isolation of distinct cell populations using magnetic sorting is a technique which has been available for nearly 3 decades, although rarely used in adult whole CNS tissue homogenate. In the current study we demonstrate that distinct cell populations can be isolated in rodents from early postnatal development through adulthood. We found this technique to be amendable to customization using commercially available membrane-targeted antibodies, allowing for cell-specific isolation across development and animal species. This technique yields RNA which can be utilized for downstream applications-including quantitative PCR and RNA sequencing-at relatively low cost and without the need for specialized equipment or fluorescently labeled cells. Adding to its utility, we demonstrate that cells can be isolated largely intact, retaining their processes, enabling analysis of extrasomatic proteins. We propose that magnetic cell sorting will prove to be a highly useful technique for the examination of cell specific CNS populations.
authors.affiliation: Department of Cell, Developmental and Integrative Biology, University of Alabama at Birmingham, Birmingham, Alabama, United States of America.
authors.forename: Leanne Melissa
authors.initials: LM
authors.lastname: Holt
authors.affiliation: Department of Cell, Developmental and Integrative Biology, University of Alabama at Birmingham, Birmingham, Alabama, United States of America.|Center for Glial Biology in Medicine, University of Alabama at Birmingham, Birmingham, Alabama, United States of America.
authors.forename: Michelle Lynne
authors.initials: ML
authors.lastname: Olsen
mesh_terms: D000818:Animals; D001253:Astrocytes / Q000378:metabolism; D015415:Biomarkers; D002540:Cerebral Cortex / Q000166:cytology* / Q000254:growth & development / Q000378:metabolism; D027342:Excitatory Amino Acid Transporter 2 / Q000032:analysis / Q000276:immunology; D015870:Gene Expression; D049452:Green Fluorescent Proteins; D018189:Immunomagnetic Separation; D051379:Mice; D008810:Mice, Inbred C57BL; D017628:Microglia / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000032:analysis*; D009474:Neurons / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D060888:Real-Time Polymerase Chain Reaction
chemical_list: D015415:Biomarkers; D027342:Excitatory Amino Acid Transporter 2; D009419:Nerve Tissue Proteins; D012333:RNA, Messenger; C493395:Slc1a2 protein, rat; D049452:Green Fluorescent Proteins