Optimization of Capillary Vibrating Sharp-Edge Spray Ionization for Native Mass Spectrometry of Triplex DNA

Area of Science:

• Analytical Chemistry
• Biochemistry
• Mass Spectrometry

Background:

• Native mass spectrometry (MS) enables the study of biomolecular structures without denaturing conditions.
• Ion production efficiency is crucial for sensitive detection of large oligonucleotides like DNA triplexes.
• Capillary vibrating sharp-edge spray ionization (cVSSI) is a novel technique for MS ionization.

Purpose of the Study:

• To investigate the impact of applied voltage and heated inlet transfer tube temperature on DNA triplex ion production using cVSSI.
• To optimize cVSSI parameters for enhanced native MS analysis of large oligonucleotides in negative-ion mode.

Main Methods:

• Utilized capillary vibrating sharp-edge spray ionization (cVSSI) for sample introduction into a mass spectrometer.
• Systematically varied applied voltage (-900 to -1500 V) and heated inlet transfer tube temperatures (250 to 450 °C).
• Analyzed the abundance of DNA triplex ions (Tri), triplex adduct ions (Tri+ad), and DNA triplex fragment ions (Tri-fr) using mass spectrometry.

Main Results:

• Medium applied voltage (-900 to -1000 V) significantly increased desired triplex ion intensities (up to ~260-fold) compared to higher voltages.
• Optimal capillary inlet temperatures between 300-400 °C maximized triplex ion abundances, with specific optima for different charge states.
• Higher temperatures (450 °C) led to decreased ion abundances, while fragment ions peaked at 400 °C due to in-source activation.

Conclusions:

• Optimal conditions for cVSSI of DNA triplexes involve medium applied voltage (~ -900 V) and inlet temperatures of 300-350 °C.
• These optimized parameters enhance the production of native DNA triplex ions, crucial for sensitive analysis.
• The findings establish a foundation for sensitive native MS of large oligonucleotides using cVSSI in negative-ion mode.