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SALSA MLPA Probemix P315 EGFR

Tumours, solid

Region: EGFR 7p11.2

General information
The SALSA MLPA Probemix P315 EGFR is a research use only (RUO) assay for the detection of deletions or duplications in the human EGFR gene. This probemix can also be used to detect the presence of EGFR c.2573T>G=p.L858R and c.2369C>T=p.T790M point mutations.

The epidermal growth factor receptor (EGFR) is a cell surface tyrosine kinase (TK) enzyme involved in controlling cell growth. Ligand-induced stimulation of EGFR followed by subsequent formation of homodimers or heterodimers with other receptors, results in the activation of downstream signalling cascades that regulate cell proliferation, differentiation, survival and DNA synthesis (Kovacs et al., 2015; Wee et al., 2017). Various genetic alterations resulting in oncogenic activity of EGFR have been described to be pivotal in tumour development and progression in multiple cancer types (Lynch et al., 2004; Bhargava et al., 2005; Brennan et al., 2013). Moreover, patients with tumours harbouring EGFR alterations tend to have a more aggressive form of the disease, and expression levels of EGFR are highly predictive of clinical outcome for cancer patients (Uribe et al., 2021).

Genetic alterations that result in oncogenic activity of EGFR in cancer include copy number amplifications and deletions, structural rearrangements of the gene, and activating mutations (Lynch et al., 2004; Bhargava et al., 2005; Brennan et al., 2013). One of the most common EGFR deletions detected in various tumour types is the EGFR deletion variant III (EGFRvIII), resulting from an in-frame deletion of exons 2-7 leading to a ligand-independent receptor (Gan et al., 2013). In addition, numerous other EGFR (exon) deletions and duplications are found in tumour biopsies. EGFR mutations cluster in the kinase domain of EGFR (exons 18-21), and cause ligand-independent activation of the receptor. Therefore, these activating mutations represent possible targets for therapeutic intervention. In this regard, certain somatic EGFR mutations as well as gene amplification in patients with non-small cell lung cancer (NSCLC) highly correlate with the clinical response to TK inhibitors. Two frequent EGFR mutations, c.2573T>G=p.L858R and c.2369C>T=p.T790M, are shown to be an important mechanism of resistance to drugs acting on the TK domain of EGFR (Uribe et al. 2021).

This SALSA MLPA probemix is not CE/FDA registered for use in diagnostic procedures. Purchase of this product includes a limited license for research purposes.

Probemix content
The SALSA MLPA Probemix P315-C2 EGFR contains 47 MLPA probes with amplification products between 124 and 472 nucleotides (nt). This includes 30 probes for the EGFR gene on 7p11.2, covering all EGFR exons except for exon 11. Furthermore, this probemix also contains 2 probes specific for the c.2369C>T= p.T790M and c.2573T>G= p.L858R EGFR mutations which will only generate a signal when the respective mutation is present. In addition, 14 reference probes are included that detect autosomal chromosomal locations, targeting relatively copy number stable regions in various cancer types. Complete probe sequences and the identity of the genes detected by the reference probes are available online (www.mrcholland.com) and in Table 3 of the Product Description.

This probemix contains nine quality control fragments generating amplification products between 64 and 105 nt: four DNA Quantity fragments (Q-fragments), two DNA Denaturation fragments (D-fragments), one Benchmark fragment, and one chromosome X and one chromosome Y-specific fragment. More information on how to interpret observations on these control fragments can be found in the MLPA General Protocol and online at www.mrcholland.com.

SALSA Binning DNA SD006
The SD006 Binning DNA provided with this probemix can be used for binning of all probes including the EGFR mutation-specific probes 22215-SP0449-L21566 for the c.2573T>G=p.L858R mutation and 17162-SP0448-L21565 for the c.2369C>T=p.T790M mutation. SD006 Binning DNA is a mixture of genomic DNA from healthy individuals and plasmid DNA that contains the target sequence detected by the above mentioned probes. Inclusion of one reaction with 5 μl SD006 Binning DNA in initial MLPA experiments is essential as it can be used to aid in data binning of the peak pattern using Coffalyser.Net software. Furthermore, Binning DNA should be included in the experiment whenever changes have been applied to the set-up of the capillary electrophoresis device (e.g. when capillaries have been renewed). Binning DNA should never be used as a reference sample in the MLPA data analysis, neither should it be used in quantification of mutation signal(s). It is strongly advised that all samples tested are extracted with the same method and derived from the same source of tissue. For further details, please consult the SD006 Binning DNA product description, available online: www.mrcholland.com. This product is for research use only (RUO).

Sample DNA
Sample DNA developed for this product:

Order Items

Probemix

Item no.
Description
Technology
Price
P315-025R
SALSA MLPA Probemix P315 EGFR – 25 rxn
€ 281.00
P315-050R
SALSA MLPA Probemix P315 EGFR – 50 rxn
€ 550.00
P315-100R
SALSA MLPA Probemix P315 EGFR – 100 rxn
€ 1075.00

Required Reagents (Sold Separately)

Item no.
Description
Technology
Price
EK1-FAM
SALSA MLPA Reagent Kit – 100 rxn – FAM
€ 341.00
EK1-Cy5
SALSA MLPA Reagent Kit – 100 rxn – Cy5
€ 341.00
EK5-FAM
SALSA MLPA Reagent Kit – 500 rxn – FAM
€ 1571.00
EK5-Cy5
SALSA MLPA Reagent Kit – 500 rxn – Cy5
€ 1571.00
EK20-FAM
SALSA MLPA Reagent Kit – 2000 rxn – FAM
€ 6037.00

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