(722d) Biophysical Analysis of Algal Decision Making during Phototactic Movement Using a 3D-Printed Microfluidic Device

Microalgae have the potential to serve as a sustainable energy source using photobioreactors and raceway ponds, yet these cultivation methods have limitations including uneven cellular distribution near the light source. By studying the decision-making abilities of the unicellular green algae species Chlamydomonas reinhardtii, researchers can investigate biological pathways regulating phototaxis (the directed migration of cells in response to light gradients) and potentially identify approaches to prevent cellular aggregation near light sources in photobioreactors. Specifically, it is unknown how migrating algae can overcome physical obstacles as they are migrating in response to a light gradient. A current hypothesis is that algae use a combination of both directed and random migration to avoid these obstacles. This work focuses on the development and use of a microfluidic device with three maze-like geometries. These mazes incorporate physical barriers to manipulate the phototactic migration of algal cells as they migrate away from a defined light source. Current approaches to study algal phototaxis rely on bulk measurements in flasks or employ unidirectional microfluidic channels that cannot interrogate changes in direction due to physical obstacles. To provide new insight into algal decision-making during phototaxis, an inexpensive approach was employed by using 3D-printed master molds to fabricate a microfluidic device. The 3D printed masters were used to replicate the microfluidic devices that were comprised of polydimethylsiloxane (PDMS) plasma-bonded to glass. Light gradients were generated using white and green penlights positioned near (10.7 mm) and far (110 mm) from the mazes to study the effect of light wavelength and intensity on phototactic fidelity. Two strains of Chlamydomonas reinhardtii were used: a wild type photophobic species (CC-124) and a mutated variant of CC-124 that lacks an eyespot (CC-4316). Cellular movement was characterized using light microscopy and a high-speed camera. Cell tracking microscopy data was processed by the ImageJ plug-in, TrackMate, to elucidate cellular decision-making during migration. Single cell tracking data confirmed that single algal cells were able to navigate around the obstructions during photophobic movement; however, this ability was strongly dependent upon the intensity of the light. Experiments found that cells were able to overcome physical barriers most frequently when exposed to low intensity green light; however, this behavior was also found to a lesser extent with white light. These studies also found that average velocity of the migrating cells did not change as a function of light intensity or wavelength, suggesting that directional sensing plays an important role during photophobic movement. These experiments demonstrate the ease in implementing a microfluidic approach and provide new insight on the decision-making abilities of green algae cells.