Fig 1.
Comprehensive view of Trackoscope: Design, interface, and tracking mechanics.
(A) The Trackoscope prototype in the inverted tracking position. The platform’s footprint is 900 cm2, with a tracking area of 325 cm2. (B) Displays a CAD model of the optics assembly. The objective lenses are interchangeable, permitting the observation of a variety of organisms. The Raspberry Pi HQ Camera alongside the Raspberry Pi Zero function as a webcam, allowing this custom digital microscope to interface with any device. (C) Demonstrates the Trackoscope user interface, featuring a live video feed, real-time tracking map, manual actuator controls, and data saving options. (D) Illustrates the flowchart of the tracking algorithm and the visualization of logic. Utilizing OpenCV’s built-in tracking, the algorithm pinpoints the organism’s location based on the user’s initial selection of a region of interest. The organism’s location informs the actuator’s movements to maintain the organism within the field of view.
Table 1.
Detailed component costs for assembling Trackoscope.
Fig 2.
Resolution and speed profiling of Trackoscope.
(A) Microscope image resolution of Stentor coeruleus with a 4x objective lens. (B) Detail resolution of Stentor with a 10x objective lens. (C) Dark field image of Spirostomum ambiguum with a 4x objective lens. (D) Image of a USAF 1951 resolution target captured with a 4X objective on the Raspberry Pi HQ microscope setup, showing an enlarged view of Group 4 and an intensity profile along the indicated blue line. This confirms resolution for Group 4, Element 6, equivalent to 28.51 Line Pairs/mm or a 35.08 μm resolution. (E) Image of a USAF 1951 resolution target with a 10x objective on the Raspberry Pi HQ microscope setup, showing an enlarged view of Group 6 and an intensity profile along the indicated blue line. This confirms resolution for Group 6, Element 6, equivalent to 114.04 Line Pairs/mm or an 8.77 μm resolution. (F) Chart of platform speeds across various micro-stepping settings, with examples of trackable organisms at those speeds.
Fig 3.
Trackoscope’s versatility in speed adaptation: Profiling rapid and slow microorganism movement.
(A) Bursaria truncatella tracked over 18 minutes. (B) Spirostomum ambiguum tracked over 12 minutes. (C) Blepharisma tracked over 10 minutes. (D) The violin plot shows the speed distributions of Spirostomum ambiguum, Bursaria truncatella, Blepharisma, and Actinosphaerium, illustrating the broad spectrum of speeds Trackoscope can handle.
Fig 4.
Behavioral diversity in microscopy: Trackoscope’s wide-ranging organism tracking capabilities.
(A) Stentor coeruleus assumes different geometries as it feeds (trumpet shape) and swims (spherical shape) over 25 minutes. While in the feeding position, different anatomical features such as the holdfast and oral pouch are also visible (S3 Video). (B) Actinosphaerium hunting Paramecium over 5.5 hours (S3 Video). Features such as axopods and the development of contractile vacuoles for digesting are visible throughout the track. (C) Deeplabcut analysis of Tardigrada as it slips on a petri dish over five seconds. The velocities and positions of the front two legs and mass center of the Tardigrada over the five seconds. (D) Binary fission of motile Blepharisma over 1.5 hours (S3 Video).
Fig 5.
Comparative movement analysis of Amoeba proteus: A Study of Crawling Versus Swimming Behaviors.
(A) Typical shape of Amoeba proteus swimming in a worm-like shape and visual of swimming setup. Track of Amoeba proteus swimming detailing speed and distance traveled over 1 hour. (B) Typical shape of Amoeba proteus crawling with multiple pseudopodia and visual of crawling setup. Track of Amoeba proteus crawling detailing speed and distance traveled over 1 hour. (C) Speed comparison of Amoeba proteus between swimming and crawling. Swimming has a higher average speed of 21 um/s while crawling has an average speed of 4 um/s. (D) Shape-changing tendencies based on the locomotion method through counting pseudopodia. While crawling, Amoeba proteus generate pseudopodia constantly while when swimming, only short pseudopodia are formed occasionally. (E) Pattern of a swimming Amoeba proteus capturing Paramecium over 1.5 hours (S4 Video). For this track, it was observed that every time the Amoeba proteus fed, it changed the general direction it was traveling in.