Label-free 3D tracking of biological micro-objects using RGB pinholes

• Published in: Japanese Journal of Applied Physics Vol. 63; no. 4; pp. 4 - 12
• Main Authors: Kim, Doyeop, Matsuzaki, Takahisa, Yoshikawa, Hiroshi Y., Kawamura, Ryuzo
• Format: Journal Article
• Language: English
• Published: Tokyo IOP Publishing 01.04.2024; Japanese Journal of Applied Physics
• Subjects: 3D-tracking; cell; Color coding; Defocusing; Digital imaging; Digital particle image velocimetry; Fluidics; Image analysis; Labels; microscopy; Microtechnology; Molecular motors; Particle image velocimetry; particles; Pinholes; Three dimensional analysis; Tracking; trajectory
• ISSN: 0021-4922; 1347-4065
• DOI: 10.35848/1347-4065/ad3969

Abstract Defocusing digital particle image velocimetry is a method developed within the field of fluidics to measure the velocities of particles within a volume through image analysis. For three-dimensional (3D) tracking, approaches utilizing defocusing techniques have been proposed to enable depthwise positional measurements. Recently, there have been suggestions to simplify this approach by employing a triple pinhole mask and to further enhance it with color filters that allow distinguish particles, even under dense conditions. This paper presents an attempt to apply the defocusing method using a triple pinhole mask with red, green, and blue (RGB) color-coding filters, demonstrating the successful label-free 3D tracking of biological micro-objects at multiple positions within a volume. The application of this method in fluidics and its potential for biological analysis are highlighted through successful examples of 3D tracking involving micro-particles influenced by motor proteins, dividing cells, and swimming microorganisms.