Kit for assessing mitochondrial toxicity
View Patent ↗Processes and methods for the simultaneous quantification of nucleic acids in diseased cells that are based on real-time PCR are provided. The real-time PCR protocol is an excellent tool for reliable quantification of in vitro drug screening and evaluation protocols to determine the efficacy of potential anti-viral agents. Quantification using these simultaneous PCR cycle threshold (Ct) detection techniques during one-step real-time RT-PCR (Applied Biosystems, CA) eliminates the variability resulting from quantification of end-point RT-PCR products. In addition, the mitochondrial toxicity assay is an added tool to assess potential side-effects for these chemotherapeutic agents.
1. A kit for assessing mitochondrial toxicity of a compound, comprising
a mixture of oligonucleotides comprising
at least one first primer set that provides a first detectable signal on the occurrence of amplification of mitochondrial nucleic acid; and
at least one second primer set that provides a second detectable signal on the occurrence of amplification of host nuclear nucleic acid;
wherein the second primer set comprises the primers of SEQ ID No. 1 and SEQ ID No. 2.
2. The kit according to claim 1 further comprising a probe of SEQ ID No. 3 comprising a reporter molecule and a quencher molecule.
3. The kit according to claim 2 , wherein the reporter molecule is FAM and the quencher molecule is TAMRA.
4. The kit according to claim 1 , wherein the first primer set comprises the primers of SEQ ID No. 19 and SEQ ID No. 20.
5. The kit according to claim 4 further comprising a probe of SEQ ID No. 21 comprising a reporter molecule and a quencher molecule.
6. The kit according to claim 5 , wherein the reporter molecule is TET and the quencher molecule is TAMRA.
7. A kit for assessing mitochondrial toxicity of a compound, comprising
a mixture of oligonucleotides comprising
at least one first primer set that provides a first detectable signal on the occurrence of amplification of mitochondrial nucleic acid; and
at least one second primer set that provides a second detectable signal on the occurrence of amplification of host nuclear nucleic acid;
wherein the first primer set comprises the primers of SEQ ID No. 19 and SEQ ID No. 20.
8. The kit according to claim 7 , wherein the second primer set comprises the primers of SEQ ID No. 1 and SEQ ID No. 2.
9. The kit according to claim 8 further comprising a probe of SEQ ID No. 3 comprising a reporter molecule and a quencher molecule.
10. The kit according to claim 9 , wherein the reporter molecule is FAM and the quencher molecule is TAMRA.
11. The kit according to claim 7 , wherein the first primer set comprises the primers of SEQ ID No. 19 and SEQ ID No. 20.
12. The kit according to claim 11 further comprising a probe of SEQ ID No. 21 comprising a reporter molecule and a quencher molecule.
13. The kit according to claim 12 , wherein the reporter molecule is TET and the quencher molecule is TAMRA.