| RRC ID |
71728
|
| 著者 |
Yasui R, Matsui A, Sekine K, Okamoto S, Taniguchi H.
|
| タイトル |
Highly Sensitive Detection of Human Pluripotent Stem Cells by Loop-Mediated Isothermal Amplification.
|
| ジャーナル |
Stem Cell Rev Rep
|
| Abstract |
For safe regenerative medicines, contaminated or remaining tumorigenic undifferentiated cells in cell-derived products must be rigorously assessed through sensitive assays. Although in vitro nucleic acid tests offer particularly sensitive tumorigenicity-associated assays, the human pluripotent stem cell (hPSC) detectability is partly constrained by the small input amount of RNA per test. To overcome this limitation, we developed reverse transcription loop-mediated isothermal amplification (RT-LAMP) assays that are highly gene specific and robust against interfering materials. LAMP could readily assay microgram order of input sample per test and detected an equivalent model of 0.00002% hiPSC contamination in a simple one-pot reaction. For the evaluation of cell-derived total RNA, RT-LAMP detected spiked-in hPSCs among hPSC-derived trilineage cells utilizing multiple pluripotency RNAs. We also developed multiplex RT-LAMP assays and further applied for in situ cell imaging, achieving specific co-staining of pluripotency proteins and RNAs. Our attempts uncovered the utility of RT-LAMP approaches for tumorigenicity-associated assays, supporting practical applications of regenerative medicine.
|
| 巻・号 |
18(8)
|
| ページ |
2995-3007
|
| 公開日 |
2022-12-1
|
| DOI |
10.1007/s12015-022-10402-3
|
| PII |
10.1007/s12015-022-10402-3
|
| PMID |
35661077
|
| PMC |
PMC9622575
|
| MeSH |
Humans
Nucleic Acid Amplification Techniques* / methods
Pluripotent Stem Cells*
RNA
Sensitivity and Specificity
|
| リソース情報 |
| ヒト・動物細胞 |
HiPS-RIKEN-2A(HPS0009)
HiPS-RIKEN-12A(HPS0029)
Nips-B2(HPS0223) |
