PurposeImmunological phenomena are a minor criteria in the modified Duke Criteria for endocarditis. Given the changes in epidemiology and diagnostics, the added value of determining these phenomena in today’s patients with suspected endocarditis is unknown.MethodsIn a retrospective cohort study of all patients with suspected endocarditis admitted to our hospital and discussed in our endocarditis team, we determined the proportion of patients classified as definite endocarditis because of either positive IgM rheumatoid factor (IgM RF), haematuria, or Roth’s spots on ophthalmology consultation. We also determined diagnostic accuracy of each of these immunological phenomena separately and combined.ResultsOf 285 patients included, 138 (48%) had definite endocarditis and at least one immunological test was performed in 222 patients (78%). Elevated IgM RF was found in 22 of 126 patients tested (17%), haematuria in 78 of 196 tested (40%) and Roth’s spots in six of 120 tested (5%). Eighteen of 138 patients with definite IE (13%) were classified as such because of a positive IgM RF, haematuria or Roth’s spots. Haematuria had the highest sensitivity: 50.5% (95% CI 40.4–60.6) and Roth’s spots the highest specificity: 98.3% (95% CI 90.8–99.9). The diagnostic accuracy results were robust in a sensitivity analysis aimed at avoiding incorporation bias.ConclusionAmong patients with a clinical suspicion of endocarditis, recommended systematic testing for immunological phenomena helped classify more patients as definite IE and is useful to confirm the diagnosis of endocarditis.Supplementary InformationThe online version contains supplementary material available at 10.1007/s15010-022-01954-0.