Application Guide - QIAxcel Advanced

50 QIAxcel Advanced Application Guide 10/2016 Discussion This pilot study showed detection of exonic deletions/duplications in both FAP and HNPCC samples using the QIAxcel Advanced System with comparable resolution and accuracy when using a sequencer. Probemix sets (MRC-Holland) for MLPA analysis have specific sizes (in bp) for the different probes in each kit, which are either reference probes or target exonic regions. This specific peak pattern is needed to differentiate the different peaks in the analysis and the pattern of peaks is specific for each analysis. We observed a deviation of 1–9 bp in our generated electropherogram peak pattern for APC with the QIAxcel ScreenGel software version 1.2 compared with the fragment size analysis using the capillary sequencer. Corrections for the size deviation are normally done at two points: within the sequencer software by applying the correct binning and then subsequently within the Coffalyser software by choosing the correct kit. The Coffalyser software is currently incompatible with the QIAxcel file format, since the Coffalyser requires .fsa file format (the output file format from the sequencer) to perform the analysis. This necessitated QIAxcel results to be analyzed manually. Nevertheless, all events in the pilot samples identified by ABI PRISM 3100 Genetic Analyzer and Coffalyser analysis were still identifiable with the QIAxcel ScreenGel analysis software after using the QIAxcel Advanced System for separation, detection and analysis. To further develop the MLPA analysis with QIAxcel Advanced System, we are currently trying to convert the output format to be compatible with the Coffalyser. Conclusions • The quality of the MLPA analysis when using the QIAxcel Advanced system is comparable with the data provided by a sequencer, both in terms of resolution and accuracy. • The QIAxcel Advanced system provides a faster alternative to current sequencer-based methods and is more cost-effective. • QIAxcel ScreenGel software can be used for MLPA analysis if the analysis is performed manually. Achieving software compatibility between QIAxcel ScreenGel and Coffalyser software would offer an even more attractive approach to MLPA analysis.

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