Abstract Background: Farming is often a family and multigenerational business. Relatedness among farmers could bias gene–environment interaction analysis. To evaluate the potential relatedness of farmers, we used data from a nested case–control study of prostate cancer conducted in the Agricultural Health Study (AHS), a prospective study of farmers in Iowa and North Carolina. Methods: We analyzed the genetic data for 25,009 SNPs (single-nucleotide polymorphisms) from 2,220 White participants to test for cryptic relatedness among these farmers. We used two software packages: (i) PLINK, to calculate inbreeding coefficients and identity-by-descent (IBD) statistics and (ii) EIGENSOFT, to perform a principal component analysis on the genetic data. Results: Inbreeding coefficients estimates and IBD statistics show that the subjects are overwhelmingly unrelated, with little potential for cryptic relatedness in these data. Conclusions: Our analysis rejects the hypothesis that individuals in the case–control study exhibit cryptic relatedness. Impact: These findings are important for all subsequent analyses of gene–environment interactions in the AHS. Cancer Epidemiol Biomarkers Prev; 23(10); 2192–5. ©2014 AACR.