Locascio attended James Madison University from 1979 to 1983 where she earned her B.Sc. in chemistry with a minor in biochemistry. In 1982, Locascio was a research assistant in the department of chemistry at West Virginia University. She attended the University of Utah from 1983 to 1986 while working as a research assistant in the department of bioengineering. Locascio completed her M.Sc. in bioengineering in 1986.
From 1986 to 1999, Locascio was a research biomedical engineer in the molecular spectroscopy and microfluidic methods group in the analytical chemistry division of the National Institute of Standards and Technology (NIST).  She received a certificate of recognition from the United States Department of Commerce in 1987, 1989, and 1990. Locascio was awarded the Department of Commerce Bronze Medal in 1991. While working at NIST, she was encouraged by her manager Willie E. May and mentor Richard Durst to pursue a doctoral degree. From 1995 to 1999, Locascio completed a Ph.D. in toxicology at the University of Maryland School of Medicine. At the University of Maryland, Katherine S. Squibb and Bruce O. Fowler, the director of the toxicology program, supported Locascios efforts to attend graduate school while also working at NIST. Her dissertation was titled Miniaturization of bioassays for analytical toxicology'. Cheng S. Lee was her doctoral advisor and Mohyee E. Eldefrawi served on her advisory committee.