The SALSA MLPA Probemix P088 Oligodendroglioma 1p-19q is an in vitro diagnostic (IVD)1
or research use only (RUO) semi-quantitative assay2
for the detection of copy number changes in chromosome arms 1p and 19q, deletions of CDKN2A
, and for the detection of four most common somatic point mutations in IDH1
(p.R132C and p.R132H) and IDH2
(p.R172K and p.R172M) in genomic DNA isolated from fresh-frozen or formalin-fixed paraffin-embedded (FFPE) human glioma specimens. P088 Oligodendroglioma 1p-19q is intended to aid in diagnosis of oligodendroglioma.
Copy number variations (CNVs) detected with P088 Oligodendroglioma 1p-19q should be confirmed with a different technique. In particular, CNVs detected by only a single probe always require confirmation by another method. Except for the four mutations mentioned above, no other mutations in IDH1
can be detected with probemix P088. For detection of rare mutations in IDH1
, it is recommended to use this assay in combination with sequence analysis.
Assay results are intended to be used in conjunction with other clinical and diagnostic findings, consistent with professional standards of practice, including confirmation by alternative methods. The results of this test should be interpreted by a clinical molecular geneticist or equivalent.
Please note that this probemix is for in vitro diagnostic (IVD) use in the countries specified at the end of this product description. In all other countries, the product is for research use only (RUO).
To be used in combination with a SALSA MLPA Reagent Kit, Coffalyser.Net analysis software, and SD079.
Oligodendrogliomas are a type of glioma (central nervous system neoplasms) that originate from the oligodendrocytes of the brain or from a glial precursor cell. The uniquely identifying and most common genomic alteration found in oligodendrogliomas is deletion of chromosomal arms 1p and 19q, either separately or combined (~70% of oligodendrogliomas) (Reifenberger et al. 2017). The WHO Classification of Tumours of the Central Nervous System guidelines recommend molecular testing of this (co-)deletion in order to distinguish oligodendrogliomas from other gliomas (Louis et al. 2016). Most oligodendrogliomas also have mutations in IDH1
or the homologous IDH2
gene. Of these mutations, p.R132H accounts for 92% and p.R132C for 4% of all IDH1
mutations, and p.R172K accounts for 64.5% and p.R172M for 19% of all IDH2 mutations (Hartmann et al. 2009).
The SALSA MLPA probemix P088-D1 Oligodendroglioma 1p-19q contains 59 MLPA probes with amplification products between 126 and 509 nucleotides (nt). This includes 19 probes for the 1p arm plus three flanking probes for 1q arm, and 12 probes for the 19q arm plus two flanking probes for 19p arm. In addition, the probemix contains four probes specific for the p.R132H and p.R132C mutations in IDH1
and for the p.R172K and p.R172M mutations in IDH2
which will only generate a signal when the mutation is present. From a dilution series using SD079, it was shown that all four IDH
mutation-specific probes provide reliable results (which can be identified by our Coffalyser.Net software) when the allelic burden is at least 12.5%. Furthermore, the probemix contains three probes for CDKN2A
and two probes for CDKN2B
. Finally, 14 reference probes are included and target relatively copy number stable regions in central nervous system tumours, especially in oligodendrogliomas. The identity of the genes detected by the reference probes is available in Table 2b of the product description.
This probemix contains nine quality control fragments generating amplification products between 64 and 121 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.mlpa.com
SALSA Binning DNA SD079:
The SD079 Binning DNA provided with this probemix can be used as Binning DNA sample for binning of four mutation-specific probes (203 nt probe 19529-L16492 IDH1
p.R132H=c.395G>A mutation, 227 nt probe 14787-L23353 IDH1
p.R132C=c.394C>T mutation, 238 nt probe 20963-L29002 IDH2
p.R172K=c.515G>A mutation and 244 nt probe 20963-L29001 IDH2
p.R172M=c.515G>T mutation). SD079 Binning DNA is a mixture of genomic DNA from healthy individuals and synthetic DNA that contains the target sequence detected by the above mentioned probes. Inclusion of one reaction with 5 μl SD079 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), as for this purpose true mutation/SNP positive patient samples or cell lines should be used. It is strongly advised to use DNA sample and reference DNA samples extracted with the same method and derived from the same source of tissue. For further details, please consult the SD079 Binning DNA product description provided. This product is for research use only (RUO), except when used in combination with a probemix for in vitro diagnostic (IVD) purpose, as specified at the end of the product description.
Sample DNA developed for this product: