An open source 16-channel fluidics system for automating sequential fluorescent in situ hybridization (FISH)-based...

Area of Science:

• Molecular Biology
• Biotechnology
• Bioengineering

Background:

• Multiplexed fluorescent in situ hybridization (FISH) enables spatial analysis of DNA/RNA targets in cells and tissues.
• Sequential hybridization workflows for multiplexed FISH are labor-intensive due to manual liquid handling, especially with increasing rounds.

Purpose of the Study:

• To develop an open-source, low-cost, and automated fluidics system for sequential multiplexed FISH imaging.
• To reduce the labor and complexity associated with multi-round FISH assays.

Main Methods:

• A fluidics module with 16 addressable, positive pressure-driven channels controlled by solenoid valves for reagent transfer.
• A controller capable of managing up to 120 solenoid valves via serial communication for automated workflow execution.
• Demonstration using 14 rounds of sequential hybridization and imaging targeting alpha satellite repeats in HeLa cells.

Main Results:

• Successful automation of fluid exchange for sequential FISH assays.
• Robust and reliable performance across multiple hybridization and imaging rounds.
• Demonstrated utility in a complex 14-round assay targeting specific DNA sequences.

Conclusions:

• The developed fluidics system effectively automates sequential multiplexed FISH workflows.
• This simple, flexible, and low-cost system can benefit researchers performing in situ assays.
• Automation reduces labor and improves the feasibility of complex, multi-round imaging experiments.