Objective <p>This study aimed to establish an immunohistochemical method for the detection of glutathione peroxidases 1-3 (GPx1-3) in healthy and colorectal cancer tissues, to perform semi-quantitative evaluation, and to assess their correlation with disease severity to explore their potential as tumourigenic markers.</p> Methods <p>Tissue samples from 58 colorectal adenocarcinomas and five healthy colons were stained for GPx1-3 and evaluated and detected by the indirect immunohistochemical method under light microscopy. Statistical analysis was performed using the chi-squared test. Histological findings were compared with data such as sex, age, tumour grade, histotype, and lymph node involvement.</p> Results <p>In healthy colon tissue, strong GPx1-3 immunoreactivity was observed in epithelial and basal cells (100%). In colorectal cancer, GPx expression was variable: GPx1 was cytoplasmically positive in 46 cases (79.31%) and negative in 12 (20.69%); GPx2 was positive in 53 (91.38%) and negative in 5 (8.62%); and GPx3 was positive in 49 (84.48%) and negative in 9 (15.52%).</p> Conclusions <p>The strong expression of GPx1-3 in normal colonic epithelium suggests their role in the pathogenesis of colorectal cancer. Their variable expression in tumour tissue suggests distinct functional roles in carcinogenesis, with potential involvement in both cancer prevention and progression. GPx1-3 may serve as promising markers for colorectal cancer. Investigation of their role may elucidate the link between oxidative stress and carcinogenesis and provide insights into new diagnostic and therapeutic strategies.</p>