On-chip HA/SSCP for the detection of hereditary haemochromatosis

Abstract

Microfluidics provides a promising tool for meeting the growing demand for high-throughput and low-cost mutation detection technology. With conventional instrumentation, this need is often addressed by the combination of the single-strand conformation polymorphism (SSCP) and heteroduplex analysis (HA) methods. This paper describes an effective microchip-based method to analyse the three most commonly tested gene mutations (C282Y, H63D, and S65C) associated with hereditary haemochromatosis by simultaneously performing microchip-based SSCP and HA, directly upon samples of polymerase chain reaction (PCR) product. We have increased the sensitivity of mutation detection considerably by adapting and combining SSCP with HA. We are able to perform the analysis within several minutes by avoiding off-chip sample preparation steps for SSCP and HA (apart from the PCR). The most important mutation in the screening of populations for this disease is the C282Y mutation and this mutation has not previously been detected with methods of HA/SSCP suitable for microchip implementation. This is, to the best of our knowledge, the first microchip-based test applying SSCP and HA for all three of the most common HFE mutations.