Four-dimensional motility tracking of biological cells by digital holographic microscopy

Abstract

Three-dimensional profiling and tracking by digital holography microscopy (DHM) provide label-free and quantitative analysis of the characteristics and dynamic processes of objects, since DHM can record real-time data for microscale objects and produce a single hologram containing all the information about their three-dimensional structures. Here, we have utilized DHM to visualize suspended microspheres and microfibers in three dimensions, and record the four-dimensional trajectories of free-swimming cells in the absence of mechanical focus adjustment. The displacement of microfibers due to interactions with cells in three spatial dimensions has been measured as a function of time at subsecond and micrometer levels in a direct and straightforward manner. It has thus been shown that DHM is a highly efficient and versatile means for quantitative tracking and analysis of cell motility.

Review of digital holographic microscopy for three-dimensional profiling and tracking

“…We recently utilized off-axis DHM and numerical autofocusing algorithm based on peak searching to image suspended polymer microspheres (9.6 μm in diameter) in suspension. 21 Figure 4(a) presents the hologram of microspheres in suspension captured by off-axis DHM. Angular spectrum method 44,45 was applied and the reconstructed amplitude image was shown in Fig.…”

“…Figures 13(f) to 13(h) show the XY, XZ, and YZ views of the 3-D trajectory, respectively. 21 In Ref. 95, a real-time hologram movie of free-swimming algae (5 to 10 μm in diameter) in seawater was recorded by in-line DHM.…”

“…We have utilized off-axis DHM to track the motility of chilomonas in a 3-D volume and time. 21 A time-lapse hologram movie of the movement of chilomonas (average diameter 10 μm) was first recorded at 30 fps. The field of view was 90 × 90 μm 2 with 464 × 464 pixels.…”

“…21 Fibers were spun onto glass coverslips from co-poly (L-gluatmic acid 4 , L-tyrosine 1 ) (PLEY) dissolved in water and crosslinked as described previously. 58 The diameter of fibers was 0.2 to 5.0 μm.…”

“…is of great interest. 21 Various approaches have been demonstrated for 3-D profiling and tracking of micro-sized objects, and optical techniques and numerical localization algorithms have been widely chosen as remarkable tools since they have the advantages of being full-field, label-free, noncontact, noninvasive, etc. Digital holographic microscopy (DHM) is an emerging technology of a new paradigm in general imaging and biomedical applications.…”