( agnostic samples andanalysed on a st...)Microfluidic analysis of multiplex,,,PCR products for the genotyping of,,,Helicobacter pylori

agnostic samples and analysed on a standard agarose gel, the assay resulted in PCR fragment patterns that were difficult to interpret in some cases.
To increase analytical precision of these studies the Agilent 2100 bioanalyzer, using Lab-on-a-chip technology was tested to replace slab gels as the tool for the electrophoretic separation and analysis of the mPCR products. First, we compared the analysis of the routinely performed combined PCRs of vacAs1/s2/cagA, of vac- Am1/m2 (data not shown) and of iceA1/2 (figure 1) on the Agilent 2100 bioanalyzer with slab gel results. The microfluidic analysis of the PCR amplicons of the H. pylori virulence genes showed and confirmed the results of our samples as imaged by slab gel electrophoresis. Especially the improved fragment resolution by the Agilent 2100 bioanalyzer (figure 1B) compared with the slab gel analysis (figure 1A) and the representation as an electropherogram (figure 1C) were helpful in differentiating DNA fragments, which differ in size by only 29 bp as with iceA1 and iceA2 (figure 1).

Next, the combined analysis of the five different alleles (vacA s1 or s2, cagA, vacA m1 or m2) was investigated (figure 2). Whereas the detection of the 5 alleles of H. pylori genotypes was sometimes difficult when imaging was performed with slab gel electrophoresis the imaging of microfluidic data showed improved analysis of distinct amplification products. The gel-like image shows the separation of vacAs2/vacAm2 in lane 1 and 4 and vacAs1/cagA/vacAm1 in lane 2 and 3 (figure 2A). The analysis resulted in calculated sizes of 98 and 101 bp for the vacAm2 amplicon and in calculated sizes of 151 and 148 bp for the vacAs2 amplicon (expected sizes: vacAm2: 102 bp and vacAs2: 150 bp, respectively). In addition to the gel like image, the result of lane 1 a
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