Microarray analysis of fluoro-gold labeled rat dopamine neurons harvested by laser capture microdissection.
The cellular heterogeneity of brain tissue presents a challenge to gene expression profiling of specific neuronal cell types. The present study employed a fluorescent neural tracer to specifically label midbrain dopamine neurons and non-dopamine cortical neurons. The labeled cells were then used to visually guide harvesting of the cells by laser capture microdissection (LCM). RNA extracted from the two populations of harvested cells was then amplified, labeled and co-hybridized to high density cDNA microarrays for two-color differential expression profiling. Many of the genes most highly enriched in the dopamine neurons were found to be genes previously known to define the dopamine neuronal phenotype. However, results from the microarray were only partially validated by quantitative RT-PCR analysis. The results indicate that LCM harvesting of specific neuronal phenotypes can be effectively guided in a complex cellular environment by specific pre-labeling of the target cell populations and underlie the importance of independent validation of microarray results.
Yao
Fayi
F
Department of Psychiatry and Behavioral Neurosciences, Wayne State University School of Medicine, Detroit, MI, USA.
Yu
Fei
F
Gong
Lijie
L
Taube
David
D
Rao
Donald D
DD
MacKenzie
Robert G
RG
D000818:Animals; D002469:Cell Separation / Q000379:methods*; D002540:Cerebral Cortex / Q000166:cytology / Q000378:metabolism; D004298:Dopamine / Q000378:metabolism*; D020869:Gene Expression Profiling / Q000379:methods*; D007834:Lasers; D008297:Male; D042282:Microdissection / Q000379:methods*; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D009474:Neurons / Q000166:cytology* / Q000378:metabolism*; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D051381:Rats; D017207:Rats, Sprague-Dawley; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D013050:Spectrometry, Fluorescence / Q000379:methods; D013194:Staining and Labeling / Q000379:methods; D013266:Stilbamidines; D018511:Systems Integration
C049774:2-hydroxy-4,4'-diamidinostilbene, methanesulfonate salt; D009419:Nerve Tissue Proteins; D013266:Stilbamidines; D004298:Dopamine