Microfluidic deposition for resolving single-molecule protein architecture and heterogeneity

Area of Science:

• Biophysics
• Nanotechnology
• Biochemistry

Background:

• Scanning probe microscopy (SPM) is crucial for nanoscale analysis of proteins.
• Sample deposition onto substrates for SPM can alter protein conformation and assembly states.
• Differential adsorption biases observations in heterogeneous protein samples.

Purpose of the Study:

• To introduce a novel single-step microfluidic spray deposition platform.
• To overcome limitations of traditional sample preparation methods in SPM.
• To enable accurate nanoscale characterization of protein morphology, mechanics, and structure.

Main Methods:

• Utilizing a microfluidic device to deposit biological solutions as subpicoliter microdroplets.
• Achieving millisecond-scale drying times, faster than protein diffusion on surfaces.
• Implementing a single-step process to transfer entire molecular content to substrates.

Main Results:

• The microfluidic spray deposition prevents surface mass transport and preserves protein assembly states.
• This method avoids conformational changes and biases in molecular populations.
• Quantitative measurements of heterogeneous systems, including protein aggregates, are enabled.

Conclusions:

• The single-step microfluidic spray deposition platform effectively preserves nanoscale molecular information.
• This technique overcomes critical limitations in SPM sample preparation for proteins.
• It facilitates accurate quantitative analysis of complex and heterogeneous biological systems.