Optimization of the Quantum-Si Platinum Single-Molecule Protein Sequencing Platform Toward Improved Complex-Matrix...

Area of Science:

• Proteomics
• Biotechnology
• Molecular Biology

Background:

• Protein sequencing is crucial for understanding biological processes and diseases.
• Next-generation protein sequencing (NGPS) offers enhanced sensitivity and precision.
• The Quantum-Si (QSi) Platinum Sequencer provides single-molecule, single-amino acid resolution.

Purpose of the Study:

• To optimize the QSi Platinum Sequencer library preparation protocol for faster sample-to-answer turnaround.
• To evaluate the performance of the optimized protocol on diverse protein samples and complex mixtures.
• To demonstrate the potential of NGPS for rapid protein characterization in complex biological systems.

Main Methods:

• Optimization of the QSi protein library preparation protocol, reducing preparation time from 32 to 10 hours.
• Application of the modified protocol to analyze single-domain antibodies and a staphylococcal enterotoxin B derivative.
• Determination of the library dilution threshold for optimal sequencing.
• Sequencing of proteins within a crude bacterial cell lysate.

Main Results:

• Sample preparation time reduced to 10 hours, enabling a sample-to-answer timeline under 24 hours.
• Successful sequencing of diverse proteins, including antibodies and enterotoxin derivatives.
• Identified a library dilution threshold of 100× for effective sequencing.
• Demonstrated successful protein sequencing in complex bacterial cell lysates.

Conclusions:

• The optimized QSi protocol significantly accelerates protein sequencing workflows.
• The technology shows promise for rapid characterization of proteins in complex biological samples.
• Further improvements in sequencing chemistry and data processing may reduce reliance on reference sequences for unknown protein identification.
• This advancement has the potential to significantly enhance protein-based diagnostic technologies.