His $7 million bequest (approximately $147.5 million in today's currency)—of which half financed the establishment of the Johns Hopkins Hospital—was the largest philanthropic gift in the history of the United States up to that time. Daniel Coit Gilman, who was inaugurated as the institution's first president on February 22, 1876, led the university to revolutionize higher education in the U.S. by integrating teaching and research. Adopting the concept of a graduate school from Germany's historic Heidelberg University, Johns Hopkins University is considered the first research university in the United States. Over the course of several decades, the university has led all U.S. universities in annual research and development expenditures. In fiscal year 2016, Johns Hopkins spent nearly $2.5 billion on research. The university has graduate campuses in Italy, China, and Washington, D.C., in addition to its main campus in Baltimore.

Johns Hopkins is organized into 10 divisions on campuses in Maryland and Washington, D.C., with international centers in Italy and China. The two undergraduate divisions, the Zanvyl Krieger School of Arts and Sciences and the Whiting School of Engineering, are located on the Homewood campus in Baltimore's Charles Village neighborhood. The medical school, nursing school, and Bloomberg School of Public Health, and Johns Hopkins Children’s Center are located on the Medical Institutions campus in East Baltimore. The university also consists of the Peabody Institute, Applied Physics Laboratory, Paul H. Nitze School of Advanced International Studies, School of Education, Carey Business School, and various other facilities.

Johns Hopkins was a founding member of the American Association of Universities. As of October 2019, 39 Nobel laureates and 1 Fields Medalist have been affiliated with Johns Hopkins. Founded in 1883, the Blue Jays men's lacrosse team has captured 44 national titles and plays in the Big Ten Conference as an affiliate member as of 2014.