Abstract
Within the nervous system, hundreds of neuronal and glial cell types have been described. Each specific cell type in the brain or spinal cord has a repertoire of cell surface molecules, or molecular determinants, through which it can be identified and characterized. Currently, robust cell identification and separation technologies require single-cell preparations to be generated while simultaneously limiting cell death and destruction of characteristic surface protein. The gentleMACS Dissociator, when used in combination with trypsin or papain-based dissociation kits, can effectively and gently dissociate brain tissue while preserving antigen epitopes and limiting cell loss. Standardized preparation of single-cell suspensions is achieved using C Tubes and optimized, preset gentleMACS Programs. Once generated, single-cell suspensions can be treated with monoclonal conjugates like Anti-Prominin-1 MicroBeads, which identify neural progenitors, or purified further using Myelin Removal Beads.
Highlights
Prepare the following solutions prior to beginning the protocol1. Hanks’ Buffered Salt Solution without divalent cations Ca2+ or Mg2+ (HBSS (w/o) 2
Within the nervous system, hundreds of neuronal and glial cell types have been described
For MACS Separations using Anti-Prominin-1 MicroBeads, P22 mouse brain was dissociated using the Neural Tissue Dissociation Kit (P)
Summary
1. Hanks’ Buffered Salt Solution without divalent cations Ca2+ or Mg2+ (HBSS (w/o) 2. HBSS, standard, i.e. with Ca2+ and Mg2+ (HBSS (w). 2. Depending on the antigen epitope of interest in subsequent applications, use either the Papain-based Neural Tissue Dissociation Kit (P) or the Trypsin-based Neural Tissue Dissociation Kit (T)
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