This kit is designed for the quantitative detection of CAR gene copy number in the genome of CAR-T/TCR-T cells prepared by using HIV-1 lentiviral vector technology.
This kit adopts the fluorescent probe method and multiplex PCR method to detect the DNA sequence related to integration or expression function on the transfer plasmid and the reference gene (RFG) in human cells, and the CAR gene copy number/cell in the sample can be calculated.
Accurate CAR copy number quantification (15-3M copies/μL) for lentiviral-modified cells
Dual detection of integrated CAR sequences & reference genes in one assay
GMP-grade precision with ≤15% CV for lot-release testing
Sample pre-treatment
Preparation of quantitative standards and PTC/NC
Reagent preparation and sample loading
Instrument run and detection
Data analysis
Assay range | 3.00×101~3.00×106copies/μL |
Limit of quantitation | 30 copies/μL |
Limit of detection | 15 copies/μL |
Precision | CV%≤15% |
| Copies/μl | Copies Log Value | Target Gene Copies Standard Curve | RFG Gene Copies Standard Curve | ||||||||
| Ct Value（1） | Ct Value（2） | CtValue（3） | Ct Mean Value | Recovery Rate | Ct Value（1） | Ct Value（2） | Ct Value（3） | Ct Mean Value | Recovery Rate | ||
| 3.00E+06 | 6.48 | 16.14 | 16.29 | 16.21 | 16.21 | 93.33% | 15.39 | 15.48 | 15.42 | 15.43 | 97.93% |
| 3.00E+05 | 5.48 | 19.27 | 19.61 | 19.43 | 19.43 | 109.50% | 18.67 | 18.79 | 18.71 | 18.72 | 101.97% |
| 3.00E+04 | 4.48 | 23.00 | 23.04 | 23.01 | 23.01 | 101.34% | 22.09 | 22.12 | 22.03 | 22.08 | 101.49% |
| 3.00E+03 | 3.48 | 26.42 | 26.53 | 26.54 | 26.54 | 100.21% | 25.43 | 25.46 | 25.45 | 25.45 | 100.45% |
| 3.00E+02 | 2.48 | 30.10 | 30.00 | 30.06 | 30.06 | 94.34% | 28.85 | 28.80 | 28.88 | 28.84 | 97.50% |
| 3.00E+01 | 1.48 | 33.41 | 33.50 | 33.27 | 33.27 | 102.29% | 32.25 | 32.18 | 32.00 | 32.14 | 100.80% |
| Amplification Efficiency | 94.4% | Amplification Efficiency | 98.82% | ||||||||
Standard curve:
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