A Double-Deck Self-Digitization Microfluidic Chip for Digital PCR.

Area of Science:

• Biotechnology
• Microfluidics
• Molecular Diagnostics

Background:

• Digital PCR (polymerase chain reaction) offers precise nucleic acid quantification.
• Existing microfluidic chips face limitations in chamber capacity and signal interference.
• Developing high-throughput and sensitive digital PCR platforms is crucial for molecular diagnostics.

Purpose of the Study:

• To present a novel double-deck microfluidic chip for enhanced digital PCR applications.
• To increase the chamber capacity and reagent volume without altering the chip's footprint.
• To eliminate fluorescence signal interference between chambers in different layers.

Main Methods:

• Fabrication of a double-deck microfluidic chip using polydimethylsiloxane (PDMS) layers sandwiched between glass substrates.
• Incorporation of over 20,000 micro-chambers per layer for droplet partitioning.
• Use of carbon black within the PDMS to prevent fluorescence crosstalk between layers.

Main Results:

• The double-deck design effectively doubles the chamber count and reagent capacity.
• Carbon black successfully blocked fluorescence interference between chip layers.
• The chip demonstrated high accuracy in absolute nucleic acid quantification over a dynamic range of 10^5.

Conclusions:

• The developed double-deck microfluidic chip provides a sensitive and high-capacity platform for digital PCR.
• Its design overcomes limitations of planar chips, enabling multiplexed detection without interference.
• This technology is poised to advance the widespread application of digital PCR in various fields.