AUTHOR=Chintalapudi Sumana R. , Djenderedjian Levon , Stiemke Andrew B. , Steinle Jena J. , Jablonski Monica M. , Morales-Tirado Vanessa M. 

TITLE=Isolation and Molecular Profiling of Primary Mouse Retinal Ganglion Cells: Comparison of Phenotypes from Healthy and Glaucomatous Retinas

JOURNAL=Frontiers in Aging Neuroscience

VOLUME=Volume 8 - 2016

YEAR=2016

URL=https://www.frontiersin.org/journals/aging-neuroscience/articles/10.3389/fnagi.2016.00093

DOI=10.3389/fnagi.2016.00093

ISSN=1663-4365

ABSTRACT=Loss of functional retinal ganglion cells (RGC) is an element of retinal degeneration that is poorly understood. This is in part due to the lack of a reliable and validated protocol for the isolation of primary RGCs. Here, we optimize a feasible, reproducible, standardized flow cytometry-based protocol for the isolation and enrichment of homogeneous RGC with the Thy1.2hiCD48negCD15negCD57neg surface phenotype. A three-step validation process was performed by (1) genomic profiling of 25-genes associated with retinal cells, (2) intracellular labeling of homogeneous sorted cells for the intracellular RGC-markers SNCG, BRN3A, TUJ1, and RBPMS; and (3) by applying the methodology on RGC from a mouse model with elevated IOP and optic nerve damage. Use of primary RGC cultures will allow for future careful assessment of important cell specific pathways in RGC.