Quantification of human neural stem cell engraftments in rat brains using ERV-3 real-time PCR.
Few sensitive and reliable methods have been available for quantifying the number of transplanted human neural stem cells (hNSC) in the animal brain. To develop an accurate method for quantifying the number of hNSC incorporated in rat brain, we performed real-time PCR on hNSC-transplanted rat brains using a target sequence for ERV-3, which is an endogenous retrovirus present with a known copy number in all human cells, but not present in rodent cells. A standard curve was developed for known amount of different mixes of hNSC and rat fibroblasts, and test samples were prepared by manually incorporating variable, predefined numbers of hNSCs into rat brains. A cerebral rat hemisphere injected with 10(7) hNSC revealed 1.125% chimerism. Moreover, a linear correlation was found between hNSC numbers injected and their concentrations in the rat brain. In conclusion, the developed quantitative ERV-3 assay enables a simple, fast, and reproducible detection and quantitation of hNSC numbers in the rat brain.
Lee
Soon-Tae
ST
Stroke and Neural Stem Cell Laboratory in Clinical Research Institute, Stem Cell Research Center, Department of Neurology, Seoul National University Hospital, Seoul, South Korea.
Chu
Kon
K
Kim
Eun-Hee
EH
Jung
Keun-Hwa
KH
Lee
Kyung-Bok
KB
Sinn
Dong-In
DI
Kim
Seung U
SU
Kim
Manho
M
Roh
Jae-Kyu
JK
D000818:Animals; D015415:Biomarkers; D001921:Brain / Q000166:cytology* / Q000821:virology; D002452:Cell Count / Q000379:methods*; D020077:Endogenous Retroviruses / Q000235:genetics*; D006801:Humans; D051379:Mice; D009474:Neurons / Q000166:cytology / Q000637:transplantation* / Q000821:virology; D051381:Rats; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012680:Sensitivity and Specificity; D033581:Stem Cell Transplantation
D015415:Biomarkers