Importance Hereditary factors play a key role in the risk of developing several cancers. Identification of a germline predisposition can have important implications for treatment decisions, risk-reducing interventions, cancer screening, and germline testing. Objective To examine the prevalence of pathogenic germline variants (PGVs) in patients with cancer using a universal testing approach compared with targeted testing based on clinical guidelines and the uptake of cascade family variant testing (FVT). Design, Setting, and Participants This prospective, multicenter cohort study assessed germline genetic alterations among patients with solid tumor cancer receiving care at Mayo Clinic cancer centers and a community practice between April 1, 2018, and March 31, 2020. Patients were not selected based on cancer type, disease stage, family history of cancer, ethnicity, or age. Exposures Germline sequencing using a greater than 80-gene next-generation sequencing platform. Main Outcomes and Measures Proportion of PGVs detected with a universal strategy compared with a guideline-directed approach and uptake of cascade FVT in families. Results A total of 2984 patients (mean [SD] age, 61.4 [12.2] years; 1582 [53.0%] male) were studied. Pathogenic germline variants were found in 397 patients (13.3%), including 282 moderate- and high-penetrance cancer susceptibility genes. Variants of uncertain significance were found in 1415 patients (47.4%). A total of 192 patients (6.4%) had incremental clinically actionable findings that would not have been detected by phenotype or family history–based testing criteria. Of those with a high-penetrance PGV, 42 patients (28.2%) had modifications in their treatment based on the finding. Only younger age of diagnosis was associated with presence of PGV. Only 70 patients (17.6%) with PGVs had family members undergoing no-cost cascade FVT. Conclusions and Relevance This prospective, multicenter cohort study found that universal multigene panel testing among patients with solid tumor cancer was associated with an increased detection of heritable variants over the predicted yield of targeted testing based on guidelines. Nearly 30% of patients with high-penetrance variants had modifications in their treatment. Uptake of cascade FVT was low despite being offered at no cost.