Immunohistochemistry (IHC) and polymerase chain reaction (PCR) and fragment separation by capillary electrophoresis represent the current clinical laboratory standard for the evaluation of microsatellite instability (MSI) status. The importance of reporting MSI status in colorectal cancer is based on its potential for guiding treatment and as a prognostic indicator. It is also used to identify patients for Lynch syndrome testing. Our aim was to evaluate pre-analytical factors, such as age of formalin-fixed and paraffin-embedded (FFPE) block, neoplastic cell percentage, mucinous component, and DNA integrity, that may influence the accuracy of MSI testing and assess the concordance between three different MSI evaluation approaches. We selected the mucinous colorectal cancer (CRC) histotype for this study as it may possibly represent an intrinsic diagnostic issue due to its low tumor cellularity. Seventy-five cases of mucinous CRC and corresponding normal colon tissue samples were retrospectively selected. MMR proteins were evaluated by IHC. After DNA quality and quantity evaluation, the Idylla™ and TapeStation 4200 platforms were adopted for the evaluation of MSI status. Seventy-three (97.3 cases were successfully analyzed by the three methodologies. Overall, the Idylla™ platform showed a concordance rate with IHC of 98.0MSS)/proficient MMR (pMMR) cases and 81.8deficient MMR (dMMR) cases. The TapeStation 4200 system showed a concordance rate with IHC of 96.0pMMR cases and 45.4dMMR cases. The concordance rates of the TapeStation 4200 system with respect to the Idylla™ platform were 98.17.8analytical factors, no significant variation in concordance rate among IHC analyses and molecular systems was observed by considering the presence of an acellular mucus cut-off textgreater50 FFPE year preparation, and DNA concentration. Conversely, the Idylla™ platform showed a significant variation in concordance rate with the IHC approach by considering a neoplastic cell percentage textgreater50p-value = 0.002), and the TapeStation 4200 system showed a significant variation in concordance rate with the IHC approach by considering a DNA integrity number (DIN) ≥4 as cut-off (p-value = 0.009). Our data pinpoint a central role of the pre-analytical phase in the diagnostic outcome of MSI testing in CRC.
- *fully automated RT—PCR
- *predictive molecular pathology