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SALSA A012 ERBB2 / HER2-neu MLPA Kit Agarose
[http://www.geneclinics.org/profiles/hht/details.html]
This MLPA kit can has been developed for the detection of the 12 amplification products with an agarose gel electrophoresis or Agilent microfluidics. Please note that the following MLPA kits are available for ERBB2 detection using a capillary sequencer: P004 ERBB2 kit: similar to A012; P105 oligodendroglioma probe mix: contains two ERBB2 probes as well as three EGFR, eight TP53, five CDKN2A and nine PTEN probes. Furthermore, the P171 Gain probe mix contains probes for many genes that are often amplified in tumours, including three ERBB2 probes and two EGFR probes. The ERBB2 gene, previously called Her2-neu, is a proto-oncogene located on human chromosome 17. It encodes the human epidermal growth factor receptor-2, which is a tyrosine kinase. Overexpression of this transmembrane cell surface protein occurs in a significant percentage of many different human cancers, among which breast, ovarian and cervical cancers1-8. Overexpression is most often the result of an increased copy number of the ERBB2 gene. Several methods are in use for the detection of ERBB2 gene amplification or ERBB2 overexpression, including immunostaining of the protein, FISH, quantitative Southern blotting and real time PCR. The MLPA technique complements these techniques as it permits detection of a gain in copy number to three copies/diploid genome in samples containing only 20-100 ng human DNA. Moreover, MLPA permits simultaneous relative quantification of up to 40 different nucleic acid sequences. Copy number changes of other genes on chromosome 17 such as TOP2A, can therefore be detected in the same reaction. Results of this MLPA kit will be similar to results obtained by quantitative Southern blots and real time PCR, as they are also performed on purified DNA. MLPA has in common with these two techniques that results are dependent on the percentage tumour cells in the sample used for DNA isolation. Like real time PCR, MLPA can be performed using DNA samples purified from formaldehyde fixed paraffin embedded tissues. MLPA probe mix A012 contains three probes that recognise different sequences of the human ERBB2 / Her2-neu gene. In addition to the ERBB2 specific probes, one probe for TOP2A, a gene located on a short distance from the ERBB2 gene as well as eight probes for single copy sequences on other chromosomes are included.
Full mix description (pdf)
IMPORTANT NOTICE: MLPA kits are sold by MRC-Holland for research purposes and to demonstrate the possibilities of the MLPA technique. This kit is not CE/FDA certified for use in diagnostic procedures. Salsa MLPA kits are supplied with all necessary buffers and enzymes. Purchase of the Salsa MLPA test kits includes a limited license to use these products for research purposes. The use of this MLPA kit requires a thermocycler with heated lid and sequence type electrophoresis equipment. Different fluorescent PCR primers are available. The MLPA technique has been first described in Nucleic Acid Research 30, e57 (2002)
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References Nederlof, P.M. et al. Poster: Detection of HER2 amplification by a new technique: MLPA, 2003.
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