Table 1.
Clinicopathologic features of colorectal cancer cases in this study (N = 336).
Fig 1.
The MSI-PCR test developed in this study.
A Workflow of microsatellite PCR analysis. The MSI-PCR test contains five poly-A mononucleotide MS markers. A marker was defined unstable in the tumour if the shift was equal to or larger than 3 bp in the major band or if new minor bands were present compared with those in the corresponding non-tumourous tissue. For the ones showing an inconclusive smearing pattern, the PCR products were re-run in a high-resolution gel to visualize the PCR bands more clearly. MSI-H: two or more of the five microsatellite markers show instability. MSI-L: one of the five microsatellite markers shows instability. MSS: no markers show instability. B Representative images of the MSI-PCR gel electrophoresis. The left panel, the MSS result. The middle panel, an MSI-H result with the major DNA products shifted by 3 bp or larger in the tumour. The sizes of the major DNA products are shown on the bottom of the gel images. The right panel, an MSI-H result with presentation of new minor DNA bands in the tumour, as compared with the non-tumourous region. T, tumour. N, non-tumourous region.
Fig 2.
Sensitivities of the MSI-PCR tests using the screening and high-resolution gel capillary electrophoresis.
Genomic DNA of the MSI (+) HCT116 cell line was mixed with that of normal FFPE tissue (A) and fresh frozen tissue (B) in various ratios for the MSI-PCR analysis. The PCR products were visualized by the QIAxcel screening (left) and high-resolution (right) gel electrophoresis for determination of the respective limit of detection (LOD). * indicates the LOD value for each MS marker.
Fig 3.
Analysis of the MMR gene variants by NGS and Sanger sequencing in the discordant case showing discordance in MSI-PCR and dMMR IHC tests.
The NGS analysis of the MMR and oncogene panel in the tumour and corresponding blood specimens were analysed by NGS. The genetic variants identified by NGS were confirmed by Sanger sequencing. VAF%, variant allele frequency.
Table 2.
Concordance between MSI-PCR and dMMR IHC analyses.