Are droplets really suitable for single-cell analysis? A case study on yeast in droplets

Area of Science:

• Biotechnology
• Cell Biology
• Microfluidics

Background:

• Single-cell analysis is vital in biotechnology, utilizing microfluidic devices like droplets for cell cultivation.
• A key assumption is that microenvironments do not harm cells, ensuring unbiased studies.
• Current assessments often overlook sub-lethal effects on cellular functions, focusing on viability.

Purpose of the Study:

• To evaluate the impact of droplet cultivation on cellular function.
• To use yeast morphology as a sensitive indicator of sub-lethal stress.
• To identify potential biases introduced by microfluidic culture conditions.

Main Methods:

• Yeast cells were cultured in flasks, large droplets, and small droplets.
• Quantitative morphological analysis was performed using the CalMorph image-analysis program.
• Cell cycle progression and bud growth were assessed as indicators of cellular function.

Main Results:

• Significant morphological differences were observed between cells cultured in flasks, large droplets, and small droplets.
• Droplet cultivation delayed the cell cycle in the G1 phase.
• Impaired mitochondrial function was suggested by delayed bud growth, and larger droplets showed more similarity to flask cultures.

Conclusions:

• Culturing cells in droplets can induce sub-lethal effects, impacting cell cycle and mitochondrial function.
• Droplet size influences cellular morphology, with larger droplets being less disruptive.
• These findings underscore the need to assess microfluidic culture conditions for accurate single-cell analysis in biotechnology.