File Download
There are no files associated with this item.
Supplementary
-
Citations:
- Appears in Collections:
Conference Paper: A comparison of high resolution melting, allele-specific priming and sanger sequencing for the detection of BRAFV600E mutation in hairy cell leukaemia from different haematological specimens
Title | A comparison of high resolution melting, allele-specific priming and sanger sequencing for the detection of BRAFV600E mutation in hairy cell leukaemia from different haematological specimens |
---|---|
Authors | |
Issue Date | 2014 |
Publisher | Blackwell Publishing Ltd. The Journal's web site is located at http://www.blackwellpublishing.com/journals/CLH |
Citation | The 27th International Symposium on Technological Innovations in Laboratory Hematology, Hague, Netherlands, 15-17 May 2014. In International Journal of Laboratory Hematology, 2014, v. 36 suppl. 1, p. 57-58, abstract no. 424 How to Cite? |
Abstract | Introduction: Recently, the BRAFV600E mutation was discovered
as a highly specific and sensitive marker for hairy cell leukaemia
(HCL) and a potential target for drug therapy. However, the quality
of clinical specimens is often suboptimal for genotypic analysis
due to a small number of hairy cells in blood and bone marrow
aspirate. This study aims to assess the usefulness of three molecular
techniques for the detection of theBRAFV600E mutation in HCL
from different types of haematological samples.
Methods: We assessed the performance of high resolution melting
(HRM), allele-specific priming (ASP) and Sanger sequencing
(SS) for BRAFV600E detection in 17 unenriched HCL samples:
blood (n=7), marrow aspirate (n=3), ethylenediaminetetraacetic
acid (EDTA)-decalcified trephine biopsy (n=2), formic acid (FA)-
decalcified trephine biopsy (n=5).
Results: In peripheral blood and marrow aspirate samples where
DNA was well preserved, all three molecular techniques showed
analysable results in all 10 HCL except for one sample in which
HRM result was not analysable. In formalin-fixed, paraffinembedded
and decalcified trephine biopsies, DNA preservation was
less optimal. High resolution melting analysis failed to produce an
analysable result in any of the seven trephine biopsies tested. The
poor performance of HRM on trephine biopsies was not related to
the method of decalcification, the length of storage or the extent of
involvement. Allele-specific priming and SS demonstrated a better
analytical ability for the mutation. The results showed that for blood
and marrow aspirate, both HRM and ASP had a very high analytical
sensitivity (1% tumour load), diagnostic sensitivity (100%) and
specificity (100%) in analysable samples. Sanger sequencing
had a lower analytical sensitivity (4% tumour load), resulting in
false-negative analysis in some cases. High resolution melting was
technically the simplest and had the shortest turn-around-time of 2
hours. In decalcified trephine biopsies, HRM was not useful, while
SS was least demanding on sample DNA quality for a successful
analysis. However, none of the three techniques showed satisfactory
diagnostic performance for trephine biopsies. Conclusions: High
resolution melting is a cost-effective technique for initial screening
of the BRAFV600E mutation in blood and marrow aspirate samples
of suspected HCL patients. Atypical cases can be confirmed by ASP
or SS. A robust detection method for this mutation on decalcified
trephine biopsies still awaits development and validation. |
Description | Poster Presentation |
Persistent Identifier | http://hdl.handle.net/10722/204401 |
ISSN | 2023 Impact Factor: 2.2 2023 SCImago Journal Rankings: 0.555 |
DC Field | Value | Language |
---|---|---|
dc.contributor.author | So, JCC | en_US |
dc.contributor.author | Chan, A | en_US |
dc.contributor.author | Chung, LP | en_US |
dc.date.accessioned | 2014-09-19T23:34:10Z | - |
dc.date.available | 2014-09-19T23:34:10Z | - |
dc.date.issued | 2014 | en_US |
dc.identifier.citation | The 27th International Symposium on Technological Innovations in Laboratory Hematology, Hague, Netherlands, 15-17 May 2014. In International Journal of Laboratory Hematology, 2014, v. 36 suppl. 1, p. 57-58, abstract no. 424 | en_US |
dc.identifier.issn | 1751-5521 | - |
dc.identifier.uri | http://hdl.handle.net/10722/204401 | - |
dc.description | Poster Presentation | - |
dc.description.abstract | Introduction: Recently, the BRAFV600E mutation was discovered as a highly specific and sensitive marker for hairy cell leukaemia (HCL) and a potential target for drug therapy. However, the quality of clinical specimens is often suboptimal for genotypic analysis due to a small number of hairy cells in blood and bone marrow aspirate. This study aims to assess the usefulness of three molecular techniques for the detection of theBRAFV600E mutation in HCL from different types of haematological samples. Methods: We assessed the performance of high resolution melting (HRM), allele-specific priming (ASP) and Sanger sequencing (SS) for BRAFV600E detection in 17 unenriched HCL samples: blood (n=7), marrow aspirate (n=3), ethylenediaminetetraacetic acid (EDTA)-decalcified trephine biopsy (n=2), formic acid (FA)- decalcified trephine biopsy (n=5). Results: In peripheral blood and marrow aspirate samples where DNA was well preserved, all three molecular techniques showed analysable results in all 10 HCL except for one sample in which HRM result was not analysable. In formalin-fixed, paraffinembedded and decalcified trephine biopsies, DNA preservation was less optimal. High resolution melting analysis failed to produce an analysable result in any of the seven trephine biopsies tested. The poor performance of HRM on trephine biopsies was not related to the method of decalcification, the length of storage or the extent of involvement. Allele-specific priming and SS demonstrated a better analytical ability for the mutation. The results showed that for blood and marrow aspirate, both HRM and ASP had a very high analytical sensitivity (1% tumour load), diagnostic sensitivity (100%) and specificity (100%) in analysable samples. Sanger sequencing had a lower analytical sensitivity (4% tumour load), resulting in false-negative analysis in some cases. High resolution melting was technically the simplest and had the shortest turn-around-time of 2 hours. In decalcified trephine biopsies, HRM was not useful, while SS was least demanding on sample DNA quality for a successful analysis. However, none of the three techniques showed satisfactory diagnostic performance for trephine biopsies. Conclusions: High resolution melting is a cost-effective technique for initial screening of the BRAFV600E mutation in blood and marrow aspirate samples of suspected HCL patients. Atypical cases can be confirmed by ASP or SS. A robust detection method for this mutation on decalcified trephine biopsies still awaits development and validation. | - |
dc.language | eng | en_US |
dc.publisher | Blackwell Publishing Ltd. The Journal's web site is located at http://www.blackwellpublishing.com/journals/CLH | - |
dc.relation.ispartof | International Journal of Laboratory Haematology | en_US |
dc.rights | The definitive version is available at www.blackwell-synergy.com | - |
dc.title | A comparison of high resolution melting, allele-specific priming and sanger sequencing for the detection of BRAFV600E mutation in hairy cell leukaemia from different haematological specimens | en_US |
dc.type | Conference_Paper | en_US |
dc.identifier.email | So, JCC: scc@pathology.hku.hk | en_US |
dc.identifier.email | Chung, LP: lpchung@hkucc.hku.hk | en_US |
dc.identifier.authority | So, JCC=rp00391 | en_US |
dc.identifier.authority | Chung, LP=rp00249 | en_US |
dc.identifier.doi | 10.1111/ijlh.12266 | - |
dc.identifier.hkuros | 237586 | en_US |
dc.identifier.volume | 36 | - |
dc.identifier.issue | suppl. 1 | - |
dc.identifier.spage | 57, abstract no. 424 | - |
dc.identifier.epage | 58, abstract no. 424 | - |
dc.publisher.place | United Kingdom | - |
dc.identifier.issnl | 1751-5521 | - |