Objectives Estimating the prevalence of drug use in the general population is important given its potential health consequences but is challenging. Self-reported surveys on drug use have inherent limitations that underestimate drug use. We evaluated the performance of linking urine drug testing with a local, representative health examination survey in estimating the prevalence of drug use in New York City (NYC). Methods We used urine drug testing from the NYC Health and Nutrition Examination Survey (NYC HANES) to estimate the prevalence of drug use (benzodiazepines, cocaine, heroin, and opioid analgesics) among the study sample and compare the findings with self-reported responses to questions about past–12-month drug use from the same survey. Results Of 1527 respondents to NYC HANES, urine drug testing was performed on 1297 (84.9%) participants who provided urine and consented to future studies. Self-reported responses gave past–12-month weighted estimates for heroin, cocaine, or any prescription drug misuse of 13.8% (95% CI, 11.6%-16.3%), for prescription drug misuse of 9.9% (95% CI, 8.1%-12.1%), and for heroin or cocaine use of 6.1% (95% CI, 4.7%-7.9%). Urine drug testing gave past–12-month weighted estimates for any drug use of 4.3% (95% CI, 3.0%-6.0%), for use of any prescription drug of 2.8% (95% CI, 1.9%-4.1%), and for heroin or cocaine use of 2.0% (95% CI, 1.2%-3.6%). Conclusion Urine drug testing provided underestimates for the prevalence of drug use at a population level compared with self-report. Researchers should use other methods to estimate the prevalence of drug use on a population level.