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. 2021 Jun 18;11:12880. doi: 10.1038/s41598-021-91974-x

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© The Author(s) 2021

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

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Endometrial dMMR/MSI tumor (case 45) with isolated loss of MSH6 expression, falsely negative by Idylla MSI assay, PCR and NGS. Hematoxylin eosin stain (a) of the tumor with classic morphology of endometrial adenocarcinoma (UCEC) with isolated loss of nuclear MSH6 (e) and preserved nuclear expression of MLH1 (b), MSH2 (c) and PMS2 (d). Idylla MSI assay (Idylla Explore software v.2.5.1294) (f) and PCR (GeneMapper v. 4.0 analysis software, Applied Biosystems). (g) called respectively 0/7 loci and 0/8 loci as MSS, resulting in MSS integrative result (Fig. 2). NGS (Python v.3.7 and Matplotlib 3.3.4 software) called 2/10 loci as unstable (Fig. 2) resulting in an overall MSS/0 score. Illustrative indel distribution plot of the FLT1, EML4 and BCL2L11 loci (h, from left to right) indicated identical number of integrated peaks (6/5/6 for respectively FLT1/ EML4/BCL2L11 loci) in tumor sample (dark blue bars) versus baseline (light blue bars) resulting in calling these loci as negative by the default mSINGS script. However, for the EML4 and BCL2L11 loci, the overall indel distribution did shift towards shorter indel lenghts as indicated by the red arrows, suggesting the presence of molecular alterations not recognized by the current parametrization of the script.