therascreen IDH1/2 RGQ PCR Kit CE
For the detection of 12 IDH1 and IDH2 mutations in glioma
The therascreen IDH1/2 RGQ PCR Kit enables detection of 12 mutations in the IDH1 and IDH2 genes using real-time PCR on the Rotor-Gene Q MDx 5plex HRM Instrument. The kit provides all necessary reagents, optimized for rapid and sensitive detection of IDH1 and IDH2 mutations.
The therascreen IDH1/2 RGQ PCR Kit CE is intended for in vitro diagnostic use in Europe.
Performance
See the therascreen IDH1/2 RGQ PCR Kit Handbook for detailed performance characteristics.
Principle
The therascreen IDH1/2 RGQ PCR Kit provides reagents to perform 9 separate amplification reactions for detection of 12 mutations:
Total reaction mixesThe Total Primers and Probe Mixes (PPM-Total) use primers and probes to amplify both mutated and wild-type target sequences. Mutation detection reaction mixesThe mutation detection primers and probe mixes combine primers and probes, to amplify both mutated and wild-type target sequences, plus an oligonucleotide, 3' blocked with the addition of a phosphate group to prevent elongation (PCR clamping), which is specific to the wild-type target sequence. When the PCR template contains the wild-type sequence, the 3'-phosphate oligonucleotide will dominate over PCR primer binding due to higher affinity. There is no or low extension by the DNA polymerase and no or low amplification is observed. When a mutated sequence is present, PCR primer binding will dominate over the 3'-phosphate oligonucleotide binding and amplification will proceed. Mutation identification reaction mixesAllele-specific amplification is achieved by ARMS (Amplification Refractory Mutation System), which exploits the ability of the DNA polymerase to distinguish between a match and a mismatch at the 3' end of a PCR primer. When the PCR primer is fully matched, the amplification proceeds with full efficiency. When the 3' base is mismatched, only low-level background amplification occurs.
Procedure
The therascreen IDH1/2 RGQ PCR Kit provides all necessary reagents for detection of 12 mutations in the IDH1 and IDH2 genes. Nine primer and probe mixes are used to amplify both mutated and wild-type target sequences. Simply add template DNA and the appropriate primer and probe mix to the ready-to-use PCR master mix, and start the reaction on the Rotor-Gene Q MDx 5plex HRM instrument using the optimized cycling program described in the kit handbook. For DNA purification, we recommend using the QIAsymphony DNA Mini Kit on the QIAsymphony SP or the QIAamp DNA FFPE Tissue Kit, following the modified protocols given in the therascreen IDH1/2 RGQ PCR Kit Handbook.
Applications
The therascreen IDH1/2 RGQ PCR Kit is an in vitro diagnostic test based on PCR technology intended for the qualitative detection of 7 mutations in the IDH1 gene and 5 mutations in the IDH2 gene, and for direct identification of 3 major mutations, in DNA extracted from formalin-fixed, paraffin-embedded (FFPE) human brain tissue.
The therascreen IDH1/2 RGQ PCR Kit is intended to be used as an aid for the classification of gliomas. |
