Hopkins, a wealthy merchant and philanthropist, was born on his family's tobacco plantation, Whitehall, in Anne Arundel County, Maryland. The second of eleven children, he was given his maternal great-grandmother’s family name, Johns. His parents were members of the Society of Friends. In 1807, when Hopkins was twelve years old, his father freed the slaves on his plantation in accordance with a decision of the meeting of the Society of Friends. In consequence, the children were required to assume more of the work of the plantation. When Hopkins was seventeen, his parents, who recognized his business aptitude, sent him to Baltimore to live and work with his uncle Gerard Hopkins in the wholesale grocery business.
Hopkins quickly learned the trade, and it was not long before he, along with three of his brothers, established Hopkins Brothers, a firm that sold provisions, including a whiskey called Hopkins’ Best, throughout the South. When he had accumulated sufficient capital, Hopkins began lending money to others, and before long he moved into banking. In time, Hopkins became the leading financier in Baltimore. He was also a canny investor. His purchase of stock in the fledgling Baltimore and Ohio Railroad came to be the base of his great wealth.
Hopkins never married; in his will, he made provisions that his fortune be used for the benefit of others. After bequests to family and servants amounting to about one million dollars, there remained approximately seven million dollars to endow the university and the hospital that bear his name.
In 1867, he selected twelve individuals to serve as trustees of a corporation called The Johns Hopkins University, whose purpose was to promote education in the state of Maryland. Likewise, a board was formed for the incorporation of The Johns Hopkins Hospital. To ensure the linking of the institutions, he named the same people to both boards that established the university and hospital.
Just before his death in 1873, Hopkins wrote a letter to the board of trustees of the hospital. In it he outlined his intent for the medical institutions: "In all your arrangements in relation to this hospital, you will bear constantly in mind that it is my wish and purpose that the institution should ultimately form a part of the medical school of that university for which I have made ample provision by my will."