title: Saturation-compensated measurements for fluorescence lifetime imaging microscopy.
abstract: Fluorophore saturation is the key factor limiting the speed and excitation range of fluorescence lifetime imaging microscopy (FLIM). For example, fluorophore saturation causes incorrect lifetime measurements when using conventional frequency-domain FLIM at high excitation powers. In this Letter, we present an analytical theoretical description of this error and present a method for compensating for this error in order to extract correct lifetime measurements in the limit of fluorophore saturation. We perform a series of simulations and experiments to validate our methods. The simulations and experiments show a 13.2× and a 2.6× increase in excitation range, respectively. The presented method is based on algorithms that can be easily applied to existing FLIM setups.
authors.forename: Yide
authors.initials: Y
authors.lastname: Zhang
authors.forename: Genevieve D
authors.initials: GD
authors.lastname: Vigil
authors.forename: Lina
authors.initials: L
authors.lastname: Cao
authors.forename: Aamir A
authors.initials: AA
authors.lastname: Khan
authors.forename: David
authors.initials: D
authors.lastname: Benirschke
authors.forename: Tahsin
authors.initials: T
authors.lastname: Ahmed
authors.forename: Patrick
authors.initials: P
authors.lastname: Fay
authors.forename: Scott S
authors.initials: SS
authors.lastname: Howard