Background-The DePaul Symptom Questionnaire (DSQ) was developed to assess the symptomatology and case definition fulfillment of individuals with myalgic encephalomyelitis (ME) and chronic fatigue syndrome (CFS). The questionnaire was recently revised to improve its psychometric properties, increase its diagnostic reliability, and assess symptoms required by case definitions. The resulting instrument was named the DSQ-2. Purpose-The current study sought to evaluate the utility and reliability of the new and revised items in the DSQ-2. Method-A cross-sectional sample of 399 adults with ME or CFS was recruited to complete the DSQ-2. Results-Descriptive analyses of the DSQ-2 suggest that the new and revised items enhance the instrument's ability to assess certain symptom domains and evaluate recent case definitions. Additionally, an exploratory factor analysis resulted in an eight-factor solution: post-exertional malaise, cognitive impairment, fever and flu, pain, sleep disruption, orthostatic intolerance, genitourinary issues, and temperature intolerance. The items within each factor demonstrated strong internal consistency reliability (Cronbach's alphas = .73-.91). Conclusion-These analyses indicate that the DSQ-2 offers a more thorough and precise understanding ME and CFS symptomology and case definition fulfillment.