Fractionation of neurons and satellite cells from human sensory ganglia in order to study herpesvirus latency.
A method is described for fractionating human trigeminal ganglia into highly purified populations of neurons and satellite cells in order to study alpha-herpesvirus latency. The method was validated by microscopy of the separated populations and by the observation that only the neuronal population, not the satellite cells, contained herpes simplex virus (HSV) DNA. The frequency of detecting HSV in neurons from ganglia was 3% (43 of 1440 neurons). HSV DNA was not detected in approximately 17,500 satellite cells. The HSV DNA genome copy number in single cells ranged from 2 to 50. These data on the frequency and cellular location of latent HSV indicate that our mechanical fractionation of cell types results in low levels of cross-contamination and provides samples from which cells infected latently can be studied at the single cell level.
Cai
Guang-Yun
GY
Department of Pediatrics, Section of Pediatric Infectious Diseases, School of Medicine, University of Colorado, Box C-227, 4200 East Ninth Avenue, Denver, CO 80262, USA.
Pizer
Lewis I
LI
Levin
Myron J
MJ
D000328:Adult; D000368:Aged; D002469:Cell Separation / Q000379:methods*; D004279:DNA, Viral / Q000032:analysis; D005260:Female; D016679:Genome, Viral; D018259:Herpesvirus 1, Human / Q000235:genetics / Q000502:physiology*; D018258:Herpesvirus 2, Human / Q000235:genetics / Q000502:physiology*; D006801:Humans; D008297:Male; D008875:Middle Aged; D009474:Neurons / Q000166:cytology / Q000821:virology; D012668:Trigeminal Ganglion / Q000166:cytology; D017735:Virus Latency / Q000502:physiology*
D004279:DNA, Viral