In-tip nanoreactors for cancer cells proteome profiling

Area of Science:

• Biochemistry
• Analytical Chemistry
• Biomedical Science

Background:

• Mass spectrometry (MS)-based proteome profiling is crucial for molecular diagnostics.
• Current protein extraction and proteolysis methods are inefficient and time-consuming.
• In-tip proteolysis offers potential for rapid sample preparation but is underdeveloped for complex samples.

Purpose of the Study:

• To develop high-performance nanoreactors encapsulated in micropipette tips for fast and multiplexed proteolysis.
• To establish an efficient in situ proteolysis platform for complex bio-samples, particularly cancer cell proteomes.
• To enhance proteomic analysis through improved sample preparation.

Main Methods:

• Nanoporous silica materials with controlled properties were synthesized and encapsulated in micropipette tips.
• The nanoreactor-filled tips were used for in situ proteolysis of cancer cell lines.
• Mass spectrometry-based bottom-up proteomic analysis was employed to evaluate performance.

Main Results:

• The developed device demonstrated high sensitivity (LOD: 0.204 ± 0.008 ng/μL), selectivity, stability, and reusability (≥10 times).
• In-tip proteolysis using nanoreactors increased protein identification by 18-52% across different cancer cell lines compared to traditional methods.
• The platform enabled comprehensive protein mapping with minimal memory effect.

Conclusions:

• Nanoreactors encapsulated in micropipette tips provide an efficient, miniaturized platform for rapid proteomic sample preparation.
• This approach significantly enhances proteome profiling, offering a promising tool for advanced biomarker discovery.
• The developed method accelerates proteolysis, making it suitable for complex biological samples.