Polyhydroxyalkanoates, or PHAs, are natural polyesters synthesized by microorganisms under unbalanced growth conditions. They have been long known for intriguing biodegradability, promising to be excellent substitutions for environmentally harming petroleum-based plastics.

 

With growing concerns about the accumulation of non-degradable plastic wastes, which can take up to 450 years to decompose, PHAs keep attracting more and more attention in research and application fields. Several imaging techniques have been used to observe the dynamics and characteristics of PHAs, including fluorescence microscopy, transmission electron microscopy (TEM), and electron cryotomography. Shape and information on the localization of PHA granules can be obtained by utilizing such systems.

 

However, the use of fluorescence and electron microscopes in PHAs research has limitations. Researchers can only acquire 2D images and/or information about cells and PHA granules at their non-native states due to the interference of the complex sample-preparing process.

 

In a recent study published in The Proceedings of the National Academy of Sciences (PNAS) [i], S.Y. Choi’s research team had a different approach to PHAs. Instead of using conventional methods such as colorimetric quantification or chromatography, her team used Tomocube’s holotomographic microscopy, to measure the intrinsic refractive index (RI) of granules in real-time. The volumes and weights of both cells and granules, granule content, and granule density could then be easily quantified at a single-cell level based on these reconstructed RI values.

 
Figure1. Quantification of individual recombinant E. coli cells accumulating PHB granules. (A) Representative 3D RI tomograms at selected x-y, y-z, and z-x planes and (B) 3D rendering images of E. coli cells with different levels of PHB accumulation.
 

Not only be able to investigate the spatiotemporal characteristics of granule formation and distribution in live bacteria, but Choi et al. could also compare those characteristics between the native and the recombinant granule-producing cells and demonstrate the distinctive features between those two.

 

This short video explains how Tomocube’s microscopy can overcome existing limitations in current research on PHA based on its revolutionary holotomography technology. Label-free, real-time, 3D-quantitative analysis in live individual cells makes the technology stand out as a powerful scientific tool in PHA research and also in other fields.

 


 

[i] Choi et al.Three-dimensional label-free visualization and quantification of polyhydroxyalkanoates in individual bacterial cell in its native state https://doi.org/10.1073/pnas.2103956118