Nitrocefin is a chromogenic substrate for β-lactamase that undergoes distinctive color change from yellow (λmax = 390 nm at pH 7.0) to red (λmax = 486 nm at pH 7.0) as the amide bond in the β-lactam ring is hydrolyzed by β-lactamase. Nitrocefin is sensitive to hydrolysis by all known lactamases produced by Gram-positive and Gram-negative bacteria. Also useful for the detection of β-lactamase patterns from bacterial cell extracts by isoelectric focusing.