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Millisecond Label-Free Single Peptide Detection and Identification Using Nanoscale Electrochromatography and
Maximillian Chibuike1,2, Chathurika Rathnayaka1,2, Suresh Shivanka1,2
1Department of Chemistry, The University of Kansas, Lawrence, Kansas 66045, United States.
Analytical Chemistry
|December 23, 2024
Summary
This study introduces a novel peptide identification technique using nanoscale electrochromatography and label-free resistive pulse sensing (RPS). The method achieves high accuracy in single-molecule peptide identification by analyzing time-of-flight and resistive pulses.
Area of Science:
- Analytical Chemistry
- Biophysics
- Nanotechnology
Background:
- Developing advanced protein identification methods is crucial for biological and medical research.
- Existing techniques often require complex sample preparation or labeling.
- Nanoscale separation and detection offer potential for high-resolution analysis of biomolecules.
Purpose of the Study:
- To develop and demonstrate a novel, label-free protein identification method.
- To investigate the use of nanoscale electrochromatography for peptide separation.
- To implement resistive pulse sensing (RPS) for label-free peptide detection and identification at the single-molecule level.
Main Methods:
- Fabrication of thermoplastic nanoelectrochromatography columns with engineered surfaces (PMMA activated by O2 plasma).
- Utilized a dual in-plane nanopore sensor for time-of-flight (ToF) elucidation.
- Performed microchip and nanochannel electrochromatography with labeled and label-free peptides, analyzing ToF, normalized RPS current amplitude, and dwell time.
Main Results:
- Peptide separation was not achieved in microchannels but was successful in nanocolumns, especially at high electric fields.
- Label-free peptide detection using the dual nanopore sensor yielded distinct resistive pulse pairs corresponding to ToF.
- Single-molecule peptide identification achieved nearly 100% classification accuracy using machine learning, with measurements <10 ms.
Conclusions:
- Nanoscale electrochromatography coupled with label-free RPS is a viable strategy for single-molecule peptide identification.
- Surface modification of nanocolumns and high electric field strengths are key for effective peptide separation.
- This technology offers a rapid and accurate method for peptide analysis without the need for molecular labels.
