therascreen PITX2 RGQ PCR Kit

For clinically reliable determination of the PITX2 methylation ratio in lymph node-positive, estrogen receptor-positive and HER2-negative high-risk breast cancer patient tissue samples

Products

The therascreen PITX2 RGQ PCR Kit is intended for in vitro diagnostic use in Europe.

therascreen PITX2 RGQ PCR Kit (8) CE

Cat. No. / ID: 873211

For 8 reactions: PITX2 RGQ PCR Master Mix, PITX2 RGQ PCR Primer Probe Mix, PITX2 RGQ PCR Reference 50, PITX2 RGQ PCR Reference Low, PITX2 RGQ PCR Negative Control and PITX2 RGQ PCR NTC

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Rotor-Gene AssayManager v2.1

Cat. No. / ID: 9024203

Software for routine testing in combination with Rotor-Gene Q and Rotor-Gene Q MDx instruments. IMPORTANT: Does not include license for use. License key must be purchased separately. Visit the Download Page for download, installation and licensing instructions.

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For up-to-date licensing information or Rotor-Gene Q instrument specific disclaimers, refer to the corresponding Rotor-Gene Q instrument user manual

Features

The first clinically validated CE-IVD DNA methylation assay to predict response to anthracycline-based chemotherapy
Provides the percent methylation ratio (PMR) to enable differentiation between patients expected to be more versus less responsive to anthracyline-based chemotherapy
Simple workflow with Sample to Insight in less than 2 working days, including automated result calling
Ready-to-use solutions and reaction mixes
Automated software using Rotor-Gene AssayManager v2.1 for quick and easy results interpretation

Product Details

The therascreen PITX2 RGQ PCR Kit is a CE-IVD kit for determination of the percent methylation ratio (PMR) in promoter 2 of the pituitary homeobox transcription factor 2 (PITX2) gene using methylation-specific real-time PCR on the Rotor-Gene Q 5-plex HRM instrument. The kit provides reagents optimized for reliable and sensitive determination of the PITX2 methylation status, and allows for a clinically validated determination of the PMR cut-off value of 12, which is predictive of patient response to anthracycline-based chemotherapy with or without endocrine therapy when used to test formalin-fixed, paraffin-embedded (FFPE) tissue samples from lymph node-positive, estrogen receptor-positive and HER2-negative high-risk breast cancer patients. Want to try this solution for the first time? Request a quote for a trial kit.

Performance

To ensure reliable and sensitive determination of the PITX2 promoter 2 methylation status, the therascreen PITX2 RGQ PCR Kit has been optimized to detect and differentiate between a methylated and unmethylated state in the target region of the PITX2 promoter 2. For samples with a low PMR, the limit of detection (LOD) is 4 and for samples with a high PMR the upper detection limit is 92, facilitating a highly reliable determination of PMR values in the range in-between. Additionally, the assay has a wide range of linearity, from PMR 5 to 50. The clinical validity of the pre-defined PMR cut-off of 12 to predict patient response to anthracycline-based chemotherapy with or without endocrine therapy was shown by using 145 patient samples with 10-year follow-up data and disease-free survival as the primary end point.

Principle

The therascreen PITX2 RGQ PCR Kit exploits the real-time PCR oligonucleotide hydrolysis principle of TaqMan probes in combination with methylation-unspecific primers. Distinction between methylated and unmethylated status of the amplified bisulfite-converted target sequence in the PITX2 promoter 2 is achieved by using the combination of a FAM-labeled probe hybridizing specifically to the methylated bisulfite-converted DNA sequence and a HEX-labeled probe specific for the unmethylated bisulfite-converted DNA sequence. The resulting FAM- and HEX-specific Ct values are then used for the automated calculation of the PMR.

Procedure

The first step is manual DNA extraction using the QIAamp DSP DNA FFPE Tissue Kit, followed by quantification of genomic DNA and then bisulfite conversion with EpiTect Fast DNA Bisulfite Kit. Bisulfite-converted DNA is then amplified by real-time PCR, with all reagents provided in the therascreen PITX2 RGQ PCR Kit and the methylation status of the PITX2 promoter 2 is determined using the Rotor-Gene Q MDx 5plex HRM instrument. Finally, the Rotor-Gene AssayManager v2.1 software is used to automatically determine the run/sample validity and calculate the PMR.

Applications

The therascreen PITX2 RGQ PCR Kit enables the determination of the percent methylation ratio (PMR) in promoter 2 of the PITX2 gene. This PMR value of PITX2 promotor 2 is predictive of the response to anthracycline-based chemotherapy regimes in lymph node-positive, estrogen receptor-positive and HER2-negative high-risk breast cancer patients. Applying PITX2 methylation as a predictive biomarker can help clinicians to select the appropriate chemotherapy regimen.

The therascreen PITX2 RGQ PCR Kit is compliant with EU IVD Directive 98/79/EC, enabling the determination of the percent methylation ratio of PITX2 promoter 2 for in vitro diagnostics.