Abstract
Phase microscopy allows stain-free imaging of transparent biological samples. One technique, using the transport of intensity equation (TIE), can be performed without dedicated hardware by simply processing pairs of images taken at known spacings within the sample. The resulting TIE images are quantitative phase maps of unstained biological samples. Therefore, spatially resolved optical path length (OPL) information can also be determined. Using low-cost, open-source hardware, we applied the TIE to living algal cells to measure their effect on OPL. We obtained OPL values that were repeatable within species and differed by distinct amounts depending on the species being measured. We suggest TIE imaging as a method of discrimination between different algal species and, potentially, non-biological materials, based on refractive index/OPL. Potential applications in biogeochemical modelling and climate sciences are suggested.
Highlights
Phytoplankton—free-living unicellular algae found throughout the world’s marine and freshwater bodies—play a pivotal role in aquatic ecosystem structure and biogeochemistry
The violin plot on the right-hand side shows the distribution of the measured relative optical path length (rOPL) values for the eight cells analysed
The rOPL values showed a clear distinction between the two species of algae tested
Summary
Phytoplankton—free-living unicellular algae found throughout the world’s marine and freshwater bodies—play a pivotal role in aquatic ecosystem structure and biogeochemistry. This primary production led to an increase in atmospheric oxygen 2 concentrations, resulting in the oxygen-rich, carbon dioxide-poor atmospheric composition This was facilitated by their role in the global carbon cycle via biomass production [2], which has led to a proportional balance of nutrients (including nitrogen, phosphorus and trace elements) between seawater and biomass—the so-called Redfield ratio [3]. The accumulation of biologically produced calcium carbonate on the seafloor (from coccolithophores as well as other calcifying marine organisms) represents the world’s largest geological sink of carbon [4] Despite these wide-ranging roles in aquatic primary production and biogeochemistry, site-specific community composition and species-specific physiological variability means that precise quantification of these roles remains poorly constrained in both marine and freshwater environments. Recent advances in measurement devices (e.g. flow cytometers) and autonomous sampling platforms (e.g. ocean gliders) have somewhat enabled this to be overcome, but remain hindered by high sensor costs, preventing replicated or networked datasets
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