Colorimetric two-dimensional photonic crystal biosensors for label-free detection of hydrogen peroxide

Hydrogen peroxide (H₂O₂) detection is highly desirable for both environment and health concerns and a number of methods have been developed to monitor H₂O₂. However, most methods are limited due to the need of sophisticated instrument and high cost. Here we report a label-free two-dimensional (2-D) photonic crystal (PhC) horseradish peroxidase/bovine serum albumin (HRP/BSA) protein composite hydrogel sensor to monitor H₂O₂. Our 2-D PhC HRP/BSA protein composite hydrogel sensor is fabricated by mild protein cross-linking of BSA and HRP with glutaraldehyde, where BSA acts as a scaffold of HRP while HRP acts as the recognition molecule. The 2-D PhC HRP/BSA protein composite hydrogel sensor selectively detects H₂O₂ since HRP can specifically decompose H₂O₂ accompanying with heme inactivation. This protein hydrogel undergoes a volume phase transition as a result of peroxidase inactivation. The inactivation-induced conformational change of HRP gives rise to a decreased cross-linking density, which enlarges the particle spacing and decreases the Debye ring of the 2-D photonic crystals (PhCs). Our optimized 2-D PhC HRP/BSA-75 protein composite hydrogel sensors show a good linear range from 8.8 × 10^-6 M to 60.6 × 10^-6 M and an excellent sensitivity with a limit of detection of 8.8 × 10^-6 M. This sensor fabrication method demonstrates the proof-of-concept for utilizing proteins with few lysine groups to develop smart PhC protein composite hydrogel sensors to determine H₂O₂ concentration in solution, which makes it possible for developing protein organogel sensors to detect gaseous H₂O₂ in the future.