Alexander Fleming married Sarah Marion McElroy, an Irish nurse, in London. Sarah was a trained nurse and they had one son together.

Alexander and Sarah Fleming had their only child, a son named Robert Fleming. Robert followed in his father's footsteps and became a medical doctor specializing in respiratory medicine.

Sarah Marion McElroy Fleming, Alexander's first wife, passed away in 1949. Their marriage lasted for over three decades and was a supportive partnership during his scientific career.

Fleming married Dr. Amalia Koutsouri-Vourekas, a Greek colleague who had been working at St. Mary's Hospital as a bacteriologist. Amalia was instrumental in helping Fleming manage his correspondence and travel arrangements, particularly after he gained international recognition. The marriage was a happy one but short due to Fleming's death two years later.

Fleming maintained strong professional relationships with his colleagues at St. Mary's Hospital and other research institutions. He valued collaboration and was known for his modesty and willingness to share his knowledge. His relationship with Almroth Wright was particularly significant, as Wright provided guidance and support early in Fleming's career. While the relationship with Florey and Chain was sometimes strained due to differing views on publicity, Fleming always acknowledged their vital role in developing penicillin into a usable drug.