Molecular Pathology BRAF Testing

I would like to access information with regard to the molecular pathology BRAF testing that you offer in your institution. I have the following questions, the answers to which I believe should be easily accessible by the laboratory and therefore shouldn’t take up too much staff time or resources.

The Trust uses Surrey Pathology Services which is based at three Surrey hospital sites: Royal Surrey County Hospital; Ashford & St Peter’s Hospitals; Frimley Hospital.

The following responses relate to Royal Surrey County Hospital NHS Foundation Trust only.

1. Do you currently offer a clinical testing service for BRAF mutation in solid tissue, specifically melanoma? (Yes, No, currently in development) Yes

2. Which methodology(ies) do you use for BRAF testing in melanoma? e.g. Real time PCR (QPCR), High resolution melting curve analysis (HRMCA), Sanger sequencing, Next Generation Sequencing (NGS), Pyrosequencing, Immunohistochemistry (IHC), Fluorescence In Situ Hybridisation (FISH), Other (please specify). If you use a specific kit I would be grateful if you could provide the name of the kit you use. Next Generation Sequencing

3. Which BRAF mutations does your methodology(ies) cover? E.g. V600E, V600K, V600D, V600R etc. All V600 mutations are detected

4. What is your current laboratory turnaround time for BRAF testing in melanoma? 7-10 days

5. What is the level of sensitivity of your BRAF methodology(ies)? SPS do not recommend testing any tissue that has less than 30% tumour, SPS sequence all regions ~500 time and do not report anything that is detected in less than 10% of these reads.

6. I understand that molecular testing in FFPE tissue can be difficult due to tissue quality and/or quantity. What would you estimate is your current failure rate for BRAF testing for melanoma? <5%

7. Approximately how many BRAF tests for melanoma would you conduct per month or year (whichever time period is most convenient for you to estimate)? ~180 per year

8. Of the BRAF tests performed for melanoma, please estimate how many (or what percentage) are found to be positive for a mutation? 35-40%

9. Of the positive tests, please estimate how many (or what percentage of the positives) are V600E? How many (or what percentage of the positives) are V600K? How many (or what percentage of the positives) are V600 all other mutations? ~ 80% V600E, 18% V600K, 2% others

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