@inbook{cbe508afee904f3aab7f62f15af3db48,
title = "Flow cytometry visualization and real-Time processing with a CMOS SPAD array and high-speed hardware implementation algorithm",
abstract = "Time-domain microfluidic fluorescence lifetime flow cytometry enables observation of fluorescence decay of particles or cells over time using time-correlated single photon counting (TCSPC). This method requires the fluorescence lifetime measured from a limited number of photons and in a short amount of time. In current implementations of the technique, the low throughput of state of the art detectors and lack of real-Time statistical analysis of the current technology, the timedomain approaches are usually coupled with off-line analysis which impedes its use in flow cell sorting, tracking and capturing. In this work, we apply a 32×32 CMOS SPAD array (MegaFrame camera) for real-Time imaging flow cytometry analysis. This technology is integrated into a 1024-beam multifocal fluorescence microscope and incorporating a microfluidic chip at the sample plane enables imaging of cell flow and identification. Furthermore, the 1.5% native pixel fill-factor of the MegaFrame camera is overcome using beamlet reprojection with <10 μW laser power at 490 nm for each beam. Novel hardware algorithms incorporating the center-of-mass method (CMM) with real-Time background subtraction and division are implemented within the firmware, allowing lossless recording of TCSPC events at a 500 kHz frame rate with 1024 histogram bins at 52 ps time resolution. Live calculation of background compensated CMM-based fluorescence lifetime is realized at a user-defined frame rate (typically 0.001 ∼ 27 kHz) for each SPAD pixel. The work in this paper considers the application of the SPAD array to confocal fluorescence lifetime imaging of multiple coincident particles flowing within a microfluidic channel. Compared to previous flow systems based on single-point detectors, the multi-beam flow system enables visualization, detection and categorization of multiple groups of cells or particles according to their fluorescence lifetime.",
keywords = "Flow cytometry, Fluorescence lifetime microscopy, Real-Time image processing, Single-photon avalanche diode",
author = "Hanning Mai and Poland, {Simon P.} and {Mattioli Della Rocca}, Francesco and Conor Treacy and Justin Aluko and Jakub Nedbal and Erdogan, {Ahmet T.} and Istvan Gyongy and Richard Walker and Ameer-Beg, {Simon M.} and Henderson, {Robert K.}",
year = "2020",
month = jan,
day = "1",
doi = "10.1117/12.2544759",
language = "English",
series = "Progress in Biomedical Optics and Imaging - Proceedings of SPIE",
publisher = "SPIE",
editor = "Farkas, {Daniel L.} and Attila Tarnok",
booktitle = "Imaging, Manipulation, and Analysis of Biomolecules, Cells, and Tissues XVIII",
address = "United States",
note = "Imaging, Manipulation, and Analysis of Biomolecules, Cells, and Tissues XVIII 2020 ; Conference date: 03-02-2020 Through 06-02-2020",
}