A methacarn fixation permits an approach that comprises multiple techniques. In this study the procedure is used to examine 100 mesenteric lymph nodes from patients with colon cancer by means of histology, immunohistochemistry (IHC), and reverse transcriptase polymerase chain reaction (RT-PCR). The evaluated nodes are found to be grossly free of metastases. The combined expression of both messenger RNA (mRNA) and protein is investigated to validate the presence of structural (cytokeratin 20, or CK20) and tumor-specific (carcinoembrionic antigen, or CEA) markers. Histological analysis shows micrometastases on 4 nodes. IHC analysis identifies isolated (CK20 and CEA positive) tumor cells on 14 other nodes. In this group, none of the nodes that are positive for CK20 IHC express the related mRNA. RT-PCR confirms the CEA IHC positivity in 50% of the cases. The double CEA IHC/RT-PCR positivity would have up-staged 33% of the pN0 cases to pN1. This approach offers a technological framework for further studies that aim to validate the clinical significance of protein/mRNA expression of tumor markers in colorectal cancer sentinel lymph nodes.
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