Single-cell analysis has become one of the main cornerstones of biotechnology, inspiring the advent of various microfluidic compartments for cell cultivation such as microwells, microtrappers, microcapillaries, and droplets. A fundamental assumption for using such microfluidic compartments is that unintended stress or harm to cells derived from the microenvironments is insignificant, which is a crucial condition for carrying out unbiased single-cell studies. Despite the significance of this assumption, simple viability or growth tests have overwhelmingly been the assay of choice for evaluating culture conditions while empirical studies on the sub-lethal effect on cellular functions have been insufficient in many cases. In this work, we assessed the effect of culturing cells in droplets on the cellular function using yeast morphology as an indicator. Quantitative morphological analysis using CalMorph, an image-analysis program, demonstrated that cells cultured in flasks, large droplets, and small droplets significantly differed morphologically. From these differences, we identified that the cell cycle was delayed in droplets during the G1 phase and during the process of bud growth likely due to the checkpoint mechanism and impaired mitochondrial function, respectively. Furthermore, comparing small and large droplets, cells cultured in large droplets were morphologically more similar to those cultured in a flask, highlighting the advantage of increasing the droplet size. These results highlight a potential source of bias in cell analysis using droplets and reinforce the significance of assessing culture conditions of microfluidic cultivation methods for specific study cases.

• PMID: 34581379
• DOI full text
• PubMed
• PubTator

Reference Type

Journal Article | Research Support, Non-U.S. Gov't

Authors

Nakagawa Y, Ohnuki S, Kondo N, Itto-Nakama K, Ghanegolmohammadi F, Isozaki A, Ohya Y, Goda K

Primary Lit For

Additional Lit For

Review For