Laser speckle jitter correction in plant fluorescence lifetime extraction

• Published in: Measurement : journal of the International Measurement Confederation 2019-02, Vol.133, p.33-40
• Main Authors: Jing, Min, Le, Jing, Hua, Deng-xin, He, Ting-yao, Zhou, Chun-yan
• Format: Article
• Language: English
• Subjects: Average relative error; Correlation analysis; Cross-correlation analysis; Displacement; Displacement vector; Fast Fourier transformations; Fluorescence; Fluorescence lifetime imaging; Inversion; Lasers; Mathematical analysis; Measurement; Vibration
• ISSN: 0263-2241; 1873-412X
• DOI: 10.1016/j.measurement.2018.09.042

•The method to minimize the influence of measurement environment on FLIM was proposed.•A cross-correlation analysis using the sequential images studied particle movement.•The fluorescence lifetime was inversely calculated through the particle displacement.•More than 18% of the non-life points achieved lifetime less than 30% relative error.•The distortion of fluorescence lifetime map caused by laser speckle jitter was solved. To minimize the influence of laser speckle jitter from a non-ideal measurement environment on fluorescence-lifetime imaging microscopy (FLIM), a cross-correlation analysis method was proposed based on the changes in displacement in the fluorescence particle inverse calculation. The displacement of each pixel in sequential images was obtained from the displacement of the laser speckle, using a fast Fourier transform cross-correlation analysis. Through the use of an accurate time-gated time resolution, FLIM was applied to investigate the test point displacement affected by the laser speckle jitter. The fluorescence intensity of the test points was then replaced based on the intensity of the displacement points. The fluorescence lifetime can finally be calculated using the iterative deconvolution algorithm. Experimental results show that the average relative error of the inverted fluorescence lifetime is less than 30%, and the effective rate of the test points without the lifetime is greater than 18%. We believe that the proposed method can provide valuable guidance toward improving the accuracy of a fluorescence- lifetime measurement.