The recent explosive growth of the field of molecular genetics has created opportunities for the development of a wide range of both diagnostic and therapeutic medical applications unimaginable just a few years ago, and promises to have a major impact on the practice of medicine into the next century. Applications of diagnostic molecular biological techniques in medical genetics make it possible not only to identify carriers of mutations associated with the more classical, clinically identified, inherited diseases such as Cystic Fibrosis, but also to potentially identify specific gene mutations in otherwise well individuals, who may then be considered to be at a higher than average risk of developing disease. The development of (hereditary) Predictive Genetics along with the utilization of molecular genetic techniques to identify acquired (somatic) genetic changes by newer quantitative PCR and/or expression microarray-based technologies, arguably represent the greatest potential area of growth in diagnostic molecular diagnostics. The earliest examples of pre-symptomatic testing have been seen in the field of Cancer Genetics, specifically the diagnosis of predisposition to Hereditary Breast or Ovarian Cancer (HBOC), and Hereditary Colon and/or Endometrial and associated cancers (Hereditary Non Polyposis Colorectal Cancer Syndrome - HNPCC), as well as the Hereditary Endocrine malignancies (MEN type 1 and 2). There has been a concomitant growth of the provision of diagnosis of acquired (somatic) genetic changes associated with disease, and the application of quantitative-PCR based approaches already allow an increasingly important and sophisticated approach to the diagnosis and medical management of disorders such as Chronic Myelogenous Leukemia as well as the Myeloproliferative disorders. In line with these more recent developments in Molecular Diagnostics a wide range of molecular techniques and specific gene tests have been instituted in the LHSC MDxL, with a special emphasis on: - genes associated with the hereditary cancer syndromes - genes associated with the hereditary peripheral neuropathies - the development of analytic methods for acquired (somatic) genetic alterations A complete list of the services provided by the MDxL can be viewed on the WebSites:MDxL at London Health Sciences Centre and/or Genetests
Current lab Technologies: The development of the sophisticated screening protocols required for the analysis of the genes noted above has involved the development of the latest methods of gene analysis utilizing DNA/RNA, derived primarily from peripheral blood samples (leukocytes), as well as from various tissues and/or cultured cells. Analytic methods in place include:
• Automated DNA extraction (Roche MagNA-Pure Compact system)
• PCR and RT-PCR
• Standard PAGE and Agarose gel electrophoresis
• Quantitative PCR (Roche)
• Liquid/Bead array (Luminex)
• DNA Heteroduplex analysis for DNA SNP’s and small I/D mutation detection primarily by DHPLC (Transgenomic WAVE system) and HRM(Roche)
• Automated DNA sequence analysis by a Siemens(VisGen)
• VNTR* and MLPA** analysis by Siemens(VisGen) system
To better handle the increasing complexity of analytic techniques applied to gene mutation screening, we have also incorporated use of a pre-PCR Robotic Sample Handling System (Beckman FX) to automate PCR set-up, especially for dhplc (or HRM)-based gene mutation screening. In addition, a further Robotic Sample Handling System (Beckman NX) has been put in place to facilitate post-PCR sample handling, specifically for DNA sequencing chemistry setup to optimize TAT’s for the exponentially increasing volume of direct DNA sequence analyses. In addition a BioRad gel reader system which enables digital storage of the results of PAGE and Agaraose gel electrophoresis analyses has been brought into use. Lab data handling utilizes an integrated customized Lab Information System (LIS) based on FileMaker Pro v10 which is run on a MAC-based LAN allowing electronic storage and retrieval of lab analytic data (e.g. dhplc data readout, MLPA assay analysis data, sequence data, etc) to expedite signing off of reports.QC Programs:
The Molecular Diagnostic Laboratory is subject to inspection and monitoring under the MOHLTC and OLA* to ensure a high level of QA and QC are maintained in the presence of good laboratory practices. QC programs include internal quality control programs as well as an external program for Molecular Diagnostic Laboratories provided by the Ontario QMPLS and the College of American Pathologists (CAP) QA/QC system for the specific tests made available by them. In addition we continue partake in specific QC programs (including sample exchange) with the Ontario Provincial Molecular Diagnostic Laboratories, as well as other Mol Dx Labs in the country to assess our proficiency in BRCA1/2 analysis. In addition collaborating with other centres across Canada (co-ordinated by Dr S. Kamel-Reid’s lab UHN, TO) to monitor the development of quantitative-PCR based approach to the management of Chronic Myelogenous Leukemia (CML). We are collaborating with the Ontario MOHLTC as they move to license all molecular genetic tests in the province. QMPLS