Microfluidic fluorescence in situ hybridization and flow cytometry (μFlowFISH).

Area of Science:

• Microfluidics
• Molecular Biology
• Environmental Science

Background:

• Identifying bacteria in complex environmental samples is crucial for understanding microbial communities and their roles in processes like bioremediation.
• Conventional methods for bacterial identification can be time-consuming and require significant sample volumes.

Purpose of the Study:

• To develop and validate an integrated microfluidic device (microFlowFISH) for rapid and sensitive bacterial identification.
• To apply the microFlowFISH device for monitoring specific bacterial species involved in metal bioremediation in contaminated groundwater.

Main Methods:

• The microFlowFISH device integrates electrophoretic loading, incubation, washing, and electrokinetic focusing for single-file cell flow.
• 16S rRNA fluorescence in situ hybridization (FISH) was used for specific bacterial labeling.
• Flow cytometry was employed for downstream detection and quantification of labeled cells.

Main Results:

• The microFlowFISH device demonstrated high labeling and detection efficiencies (74-97%) with cultured bacteria and environmental isolates.
• Results from microFlowFISH showed excellent agreement with conventional flow cytometry.
• The device successfully monitored low cell numbers (100-200 cells) of specific bacteria in contaminated groundwater samples.

Conclusions:

• The microFlowFISH offers an automated and sensitive platform for quantitative bacterial detection in complex samples.
• This technology is well-suited for analyzing precious samples with low microbial cell counts, such as those from extreme environments or bioremediation sites.