Real-time imaging of autofluorescence NAD(P)H in single human neutrophils

Real-time detection of NAD(P)H is particularly important for understanding physiological activities of neutrophils. We scrutinize the performance of weak light detection systems with electron multiplying CCDs (EMCCDs) with regard to the feasibility of valid investigations by autofluorescence NAD(P)H in single human neutrophils. The low-noise amplification facility of EMCCDs is indeed just adequate to permit detection at an irradiation level where neither quenching nor phototoxic effects occur. For demonstration, a neutrophil respiratory burst was triggered and observed in real time. Our low-intensity detection system fulfills all requirements for real-time investigations at high spatiotemporal resolution in the field of neutrophil physiology and pathology.