Chiral Sum Frequency Generation Spectroscopy Detects Double-Helix DNA at Interfaces.

Area of Science:

• Surface science
• Spectroscopy
• Biotechnology

Background:

• DNA immobilization is crucial for DNA-based technologies.
• Understanding DNA structure-function at interfaces is essential.
• Chiral sum frequency generation (SFG) spectroscopy has potential for probing DNA.

Purpose of the Study:

• To develop and compare three DNA immobilization methods for chiral SFG spectroscopy.
• To systematically investigate the chiral and achiral SFG responses of single-stranded and double-stranded DNA.
• To explore new applications of chiral SFG in DNA analysis.

Main Methods:

• Developed three DNA immobilization techniques: 'anchor' (covalent), 'island' (dropcasting), and 'buoy' (air-water interface).
• Utilized chiral and achiral sum frequency generation (SFG) spectroscopy to analyze C-H stretching spectra.
• Employed heterodyne detection for enhanced chiral SFG analysis.

Main Results:

• Chiral SFG showed distinct responses for double-stranded DNA (prominent signals) versus single-stranded DNA (null response), enabling label-free hybridization detection.
• Heterodyne chiral SFG distinguished between native (D-) and non-native (L-) DNA double helices.
• Chiral SFG detected nucleobase aromatic C-H stretching modes upon hybridization and identified various DNA secondary structures beyond the canonical double helix.

Conclusions:

• The three developed immobilization methods are compatible with chiral SFG spectroscopy for DNA analysis.
• Chiral SFG offers a versatile, label-free tool for detecting DNA hybridization, distinguishing helix handedness, and characterizing DNA secondary structures.
• This work establishes a foundation for advanced chiral SFG applications in fundamental research and biotechnology.