APIS MET Alterations Kit
The APIS MET Alterations Kit is a unique real-time PCR (RT-qPCR) assay simultaneously detecting both MET exon 14 skipping mutation (METΔex14) and MET gene overexpression, using RNA extracted from FFPE non-small cell lung cancer (NSCLC) tissue or liquid biopsy (cfRNA).
Alterations detected by the APIS MET Alterations Kit
The APIS MET Alterations Kit is designed to detect the relative expression of the wild type cMET transcript, and the RNA product of mutations in MET that lead to the exon-skipping (deletion) of exon 14 (METΔex14 amplicon).
Figure 1. Alterations detected by the APIS MET Alterations Kit (A) MET overexpression (cMET) (B) MET exon 14 skipping mutation (METΔex14).
APIS METΔex14 Assay Sensitivity (Limit of Detection)
The Limit of Detection (LoD) of METΔex14 assay was estimated by testing synthetic RNA fragments encoding METΔex14 at varying concentrations, in a background of 10ng healthy lung total RNA (containing approx. 2100 copies of the wild type transcript).
The LoD was defined as the lowest number of METΔex14 copies with a 100% detection rate (8/8 replicates).
The LoD for METΔex14 was established at 25 copies (1.17% MAF).
Table 1. LoD estimation for the METΔex14. The estimated mutant allele frequency (%MAF) for the deletion for each copy number is indicated. LoD of 25 copies shown in bold.
Limit of Blank (LoB): Determined by testing negative RNA samples derived from cell-line, and total human lung RNA. For each target, the overall rate of correct sample interpretation was ≥98%.
Detection of Relative cMET Expression
The APIS MET Alterations Kit allows for the detection of cMET relative gene expression. The expression of cMET was measured in three cell lines with pre-characterised transcript numbers (Figure 2).
Figure 2. cMET ΔCt values from three cMET characterised cell lines, MCF-7, H596 and Hs746T plotted against Log10 nTPM (normalised transcripts per million). Hs746T harbours both amplified cMET and the METΔex14 mutation, while H596 and MCF-7 are negative for MET overexpression. Results generated using QS5™ Dx instrument.
APIS MET Alterations Kit Dynamic Range
The dynamic range of cMET and the two reference genes (IPO8 and UBXN4) was determined using a dilution series of healthy lung RNA. Synthetic RNA in a wild type background of ~2200 copies was used to determine the dynamic range of METΔex14.
Table 2. Linearity analysis of each target in the APIS MET Alterations Kit.
The APIS MET Alterations Kit contains two reference genes which allow for normalisation of assay input and monitor for assay set up. Positive and Negative Controls (PC and NTC) which monitor for both assay set up and reagent performance are also supplied with the kit.
APIS MET Alterations Kit Specifications
APIS MET Alterations Kit
The APIS MET Alterations Kit is available as a research use only (RUO) product. Not applicable for use in diagnostic procedures.
For more information on this product please contact us and select ‘MET Alterations Kit’.
Ordering Information
| Product Name | Test Type | Kit Size | Catalogue Number | Price |
|---|---|---|---|---|
| APIS MET Alterations Kit | RUO | 24 samples in duplicate plus controls | 03001 | Available upon request |
To order the APIS MET Alterations Kit or to learn more about how our assay can elevate your lung cancer research capabilities, please contact us below.