A 3D-printed microfluidic gradient concentration chip for rapid antibiotic-susceptibility testing

The rise of antibiotic resistance as one of the most serious global public health threats has necessitated the timely clinical diagnosis and precise treatment of deadly bacterial infections. To identify which types and doses of antibiotics remain effective for fighting against multi-drug-resistant pathogens, the development of rapid and accurate antibiotic-susceptibility testing (AST) is of primary importance. Conventional methods for AST in well-plate formats with disk diffusion or broth dilution are both labor-intensive and operationally tedious. The microfluidic chip provides a versatile tool for evaluating bacterial AST and resistant behaviors. In this paper, we develop an operationally simple, 3D-printed microfluidic chip for AST which automatically deploys antibiotic concentration gradients and fluorescence intensity-based reporting to ideally reduce the report time for AST to within 5 h. By harnessing a commercially available, digital light processing (DLP) 3D printing method that offers a rapid, high-precision microfluidic chip-manufacturing capability, we design and realize the accurate generation of on-chip antibiotic concentration gradients based on flow resistance and diffusion mechanisms. We further demonstrate the employment of the microfluidic chip for the AST of E. coli to representative clinical antibiotics of three classes: ampicillin, chloramphenicol, and kanamycin. The determined minimum inhibitory concentration values are comparable to those reported by conventional well-plate methods. Our proposed method demonstrates a promising approach for realizing robust, convenient, and automatable AST of clinical bacterial pathogens.