Monitoring and manipulation of the pH of single cells using infrared spectromicroscopy and a molecular switch.

BACKGROUND: The pH of a biological system is a crucial determinant of the structures and reactivity of its components and cellular homeostasis of H(+) is critical for cell viability. Control and monitoring of cellular acidity are highly desirable for the purpose of studying biochemical processes in vivo. METHODS: The effect of photolysis of a caged strong acid, the ester 1-(2-nitrophenyl)-ethylhexadecyl sulfonate (HDNS) is used to cause a controlled drop in pH in single cells. An isolated cell is selected under the IR microscope, irradiated with near-UV light and monitored by FTIR. RESULTS: We demonstrate the use of FTIR spectromicroscopy to monitor light-induced acidification of the cellular medium by measuring the increased concentration of CO2 and corresponding decrease of HCO3(-) in the cell and in the surrounding medium. CONCLUSIONS: We have demonstrated a method to control and accurately monitor the changes in pH of a cellular system by coupling a caged proton-releasing agent with FTIR spectromicroscopy detection. The overall implementation of photolysis and spectroscopic detection in a microscope optical configuration ensures single cell selectivity in both acidification and monitoring. We show the viability of monitoring of pH changes by FTIR spectromicroscopy with sensitivity comparable to that of glass electrodes, better than the existing methods for determining cell pH. GENERAL SIGNIFICANCE: Reporting the effect of small variations of cellular acidity provides a major improvement in the understanding of the interplay between molecular properties as assessed in vitro and cell physiology.