Label‐Free Visualization of Carbapenemase Activity in Living Bacteria.

Evaluating enzyme activity intracellularly on natural substrates is a significant experimental challenge in biomedical research. We report a label‐free method for real‐time monitoring of the catalytic behavior of class A, B, and D carbapenemases in live bacteria based on measurement of heat changes. By this means, novel biphasic kinetics for class D OXA‐48 with imipenem as substrate is revealed, providing a new approach to detect OXA‐48‐like producers. This in‐cell calorimetry approach offers major advantages in the rapid screening (10 min) of carbapenemase‐producing Enterobacteriaceae from 142 clinical bacterial isolates, with superior sensitivity (97 %) and excellent specificity (100 %) compared to conventional methods. As a general, label‐free method for the study of living cells, this protocol has potential for application to a wider range and variety of cellular components and physiological processes. Heat of the moment: A label‐free method for real‐time monitoring of carbapenemase activity in live bacteria is based on measurement of heat changes induced by hydrolysis of the substrate imipenem. This in‐cell calorimetry approach offers major advantages in rapid screening of carbapenemase‐producing Enterobacteriaceae in clinical bacterial isolates, with superior sensitivity and excellent specificity compared to conventional methods. [ABSTRACT FROM AUTHOR]