Comparing two diagnostic laboratory tests for Williams syndrome: fluorescent in situ hybridization versus multiplex ligation-dependent probe amplification.

Most people with Williams syndrome (WS) have a heterozygous 1.55 Mb deletion on chromosome 7q11.23. For diagnostic purposes, fluorescence in situ hybridisation (FISH) with commercial FISH probes is commonly used to detect this deletion. We investigated whether multiplex ligation-dependent probe amplification (MLPA) is a reliable alternative for FISH. The MLPA kit (SALSA P029) contains probes for eight genes in the WS critical region: FKBP6, FZD9, TBL2, STX1A, ELN, LIMK1, RFC2, and CYLN2. The experimental FISH assay that was used consists of four probes covering the WS critical region. A total number of 63 patients was tested; in 53 patients, a deletion was detected both with FISH and MLPA(P029), in 10 patients both techniques failed to demonstrate a deletion. In only one patient, a deletion was detected which was not previously detected by two commercial FISH probes. This patient appeared to carry a small, atypical deletion. We conclude that MLPA is a reliable technique to detect WS. Compared with FISH, MLPA is less time consuming and has the possibility to detect also smaller, atypical deletions and duplications in the WS critical region.

Diagnostic DHPLC Quality Assurance (DDQA): a collaborative approach to the generation of validated and standardized methods for DHPLC-based mutation screening in clinical genetics laboratories.

Genetic testing in a clinical diagnostic environment must be subject to rigorous quality control procedures, in order to ensure consistency and accuracy of results. Denaturing high performance liquid chromatography (DHPLC) has become a standard prescreening tool for mutation detection, offering very high efficiency and sensitivity of detection. Despite the relatively simple software-assisted assay setup, DHPLC is a complex assay, and quality control is reliant on ensuring optimal instrument performance, excellent assay design and validation, and sufficient user training and proficiency to interpret results. We describe here a unique collaborative effort by a group of diagnostic clinical genetics laboratories with DHPLC expertise who, together with the manufacturer of one of the most widely used DHPLC platforms, have generated standard operating procedures (SOPs) for instrument operation and maintenance, and for mutation detection by DHPLC. We also describe the validation of a disease-specific SOP for DHPLC assisted mutation screening of the MECP2 gene associated with Rett syndrome. The proposed SOP was validated, and used independently in two laboratories to introduce MECP2 testing. In addition, we provide empirically derived normal ranges for the WAVE System Mutation Standards, which are essential for optimal instrument performance. This effort was initiated to try to standardize DHPLC-based mutation screening procedures across laboratories, and so increase the overall quality of this testing method. This endeavor will thus save each laboratory from having to generate SOPs on their own, which is a lengthy and laborious task. In this respect, we define "generic" SOPs as procedures that are easily adaptable to the individual laboratories' quality systems.