A Carbon Monoxide Sensing Film Based on Hemoglobin Allostery

Abstract

Carbon monoxide (CO) is a gaseous mediator, which is generated via anenzymatic reaction of heme oxygenase, and it plays physiological roles to regulate cellular respiration and blood flow in the liver. The concentration and distribution of CO molecules in the living body is unknown owing to a lack of a suitable technique for measuring them in vivo. A needle-type CO sensor has been used for bioinstrumentation, but it is inappropriate for implantation in vivo and long-term monitoring. We developed a CO sensor sheet based on hemoglobin (Hb) allostery, as Hb undergoes a conformational change on CO binding. Hb was extracted from mice blood and mixed with agarose gel with a reducer to stabilize deoxy-Hb in the gel. CO-releasing molecules (CORM) were used to mimic CO-generating tissue, and the sensitivity of the Hb gel could be regulated by Hb concentration. We defined the CO–Hb index, an absorbance ratio at 539 and 557 nm, to estimate the accumulation of captured CO in the gel. It correlatively increased with CORM dose, indicating that gel-embedded Hb underwent a conformational change on CO binding, thereby acting as a CO sensor. We subsequently used the Hb-sensor sheet for two-dimensional imaging of CO distribution. CORM-containing gels with different sizes and doses were layered on this sheet. Size- and dose-dependent CO distribution was visualized by scanning the CO-Hb index in the sheet. Our Hb-based CO sensor sheet is composed of biocompatible materials and can be applied to detect low-level CO sources in the living body.