Mobility Analysis of Proteins by Charge Reduction in a Bipolar Electrospray Source

Area of Science:

• Analytical Chemistry
• Biophysics
• Mass Spectrometry

Background:

• Analyzing large biopolymers like proteins requires reducing their charge for accurate electrical mobility measurements.
• Existing methods for charge reduction can be complex or involve radioactive sources.

Purpose of the Study:

• To develop a new, non-radioactive method for reducing the charge of large electrosprayed biopolymers.
• To improve the analysis of proteins by achieving predominantly singly charged ions.

Main Methods:

• Combining positive aqueous electrospray (ESI) of proteins with negative methanolic ESI in a single chamber.
• Utilizing a grounded metallic grid to electrostatically decouple the two sprays and reduce ion charge.
• Employing a triethylammonium formate buffer to minimize droplet size.
• Optimizing capillary tip size for the positive emitter to stabilize the aqueous spray.

Main Results:

• Achieved substantial charge reduction on multiply charged protein cations.
• Obtained protein peaks with narrow widths, comparable to previous methods.
• Demonstrated control over the charge state distribution, from high natural charges to predominantly singly charged ions.
• Successfully analyzed large proteins, including immunoglobulin G (approximately 150 kDa).

Conclusions:

• The developed dual electrospray method offers an effective and practical approach for charge reduction of large biopolymers.
• This technique facilitates the analysis of large proteins by electrical mobility, overcoming limitations of previous methods.