Tracking and Analysis of FUCCI-Labeled Cells Based on Particle Filters and Time-to-Event Analysis Kenji Fujimoto 1 , Shigeto Seno 1* , Hironori Shigeta 1 , Tomohiro Mashita 1, 2 , Masaru Ishii 3, 4 , Hideo Matsuda 1 1 Graduate School of Information Science and Technology, Osaka University, Suita, Osaka, Japan. 2 Cybermedia Center, Osaka University, Toyonaka, Osaka, Japan. 3 Graduate School of Medicine, Osaka University, Suita, Osaka, Japan. 4 Graduate School of Frontier Bioscience, Osaka University, Suita, Osaka, Japan. * Corresponding author. Tel.: +81-6-6879-4391; email: senoo@ist.osaka-u.ac.jp Manuscript submitted November 30, 2019; accepted March 4, 2020. Abstract: FUCCI (fluorescent ubiquitination-based cell cycle indicator) is a fluorescent probe used to visualize the cell cycle progression of individual cells using fluorescent proteins of different colors. Because the cell cycle is related to biological processes such as proliferation of cancer cells, analysis of imaging data visualized using FUCCI is extremely important. This paper proposes a method for spatiotemporal tracking and analysis of FUCCI-labeled cells from time-lapse videos. To address the color transition of the FUCCI-labeled cell with the cell cycle progression, the proposed method simultaneously estimates the location and the cell cycle phase of the target cell. Furthermore, to analyze the cell phase transition, this paper proposes to apply multistate time-to-event analysis to the information obtained through our tracking method. This paper demonstrates the usefulness of our method with application to FUCCI-labeled HuH7 cells (human hepatocellular carcinoma cell line). Key words: Cell cycle, cell tracking, FUCCI, time-to-event analysis, particle filter. 1. Introduction Recent advances in bioimaging technology have enabled the visualization of various biological processes, and automated systems for data acquisition have enabled high-throughput imaging. High-content screening (HCS), an imaging-based multi-parametric analysis method, is used in biological research and drug discoveries [1]. Such advances have been producing enormous amounts of data. Therefore, the development of algorithms for image quantification and analysis is an urgent issue. We focus on cell cycle analysis of cells visualized using fluorescent ubiquitination-based cell cycle indicator (FUCCI) [2]. The cell cycle is a sequence of events in a cell, which starts when a cell is produced from its parent cell’s division and finishes when the cell divides. The cell cycle is composed of four phases, i.e., gap 1 (G1), synthesis (S), gap 2 (G2), and mitosis (M). In the S phase, cells replicate their DNA to prepare for cell division in the M phase. The G1 and G2 phases are devoted to cell growth. FUCCI is a kind of fluorescent probe used to visualize phases in the cell cycle by staining cell nuclei with different fluorescence proteins according to the phase. Fig. 1 (a) shows FUCCI-labeled HuH7 cells (human hepatocellular carcinoma cell line) superimposed with red and green fluorescence on a corresponding differential interference contrast (DIC) microscopy image (gray). FUCCI cells emit red fluorescence in the G1 phase, International Journal of Bioscience, Biochemistry and Bioinformatics 94 Volume 10, Number 2, April 2020 doi: 10.17706/ijbbb.2020.10.2.94-109