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Updated: Apr 2, 2026
Label-Free Quantitative Proteomics Workflow for Discovery-Driven Host-Pathogen Interactions
Published on: October 20, 2020
Microfluidics-enabled proteomic profiling reveal iron-driven immune evasion by an antimicrobial-resistant pathogen
Chikim Nguyen1, Chelsea Reitzel2, Arjun Sukumaran2
1Autonomous Matter Department, AMOLF, 1098 XG Amsterdam, the Netherlands.
Abstract:
Dissecting host-pathogen interactions is challenging due to heterogeneous co-cultures and limited separation methods. Here, we developed a label-free microfluidic chip enabling reproducible separation of Klebsiella pneumoniae and murine macrophages during co-culture for high-resolution proteomic analysis. Using an optimized 1.4 μm filter, the platform preserved cell viability while improving host protein identification and enriching immune-associated proteins compared to traditional scraping and supernatant collection. Chip-isolated non-phagocytosed bacteria displayed distinct proteome profiles, including reduced metabolic enzymes and increased biosynthetic and iron-binding proteins. Iron-associated proteins were uniquely enriched in this population, and functional assays confirmed that iron promotes macrophage evasion and bacterial survival. Together, these results establish a microfluidic-proteomic workflow for resolving complex host-pathogen dynamics and propose an iron-dependent mechanism of immune evasion. This approach reduces sample handling and cross-contamination while preserving cellular structure, providing a powerful framework for studying infection biology and identifying therapeutic targets.
Insights
A novel microfluidic chip effectively separates host cells and bacteria, revealing how iron aids immune evasion. This breakthrough aids infection biology research and therapeutic target identification.
Area of Science:
- Microfluidics
- Proteomics
- Infectious Disease Biology
Background:
- Studying host-pathogen interactions is complex due to mixed cell populations.
- Existing separation methods limit high-resolution analysis and cell viability.
Purpose of the Study:
- To develop a label-free microfluidic chip for separating bacteria and host cells.
- To enable high-resolution proteomic analysis of host-pathogen interactions.
- To investigate bacterial immune evasion mechanisms.
Main Methods:
- Fabrication of a microfluidic chip with an optimized 1.4 μm filter.
- Co-culture of Klebsiella pneumoniae and murine macrophages on the chip.
- Label-free separation of bacteria and host cells.
- Proteomic analysis of separated bacterial populations.
- Functional assays to assess iron's role in immune evasion.
Main Results:
- The microfluidic chip enabled reproducible separation while preserving cell viability.
- Improved host protein identification and enrichment of immune-associated proteins compared to traditional methods.
- Chip-isolated bacteria showed distinct proteomes with altered metabolic and iron-binding proteins.
- Iron was found to promote bacterial survival and macrophage evasion.
Conclusions:
- The microfluidic-proteomic workflow offers a powerful tool for dissecting host-pathogen dynamics.
- An iron-dependent mechanism contributes to bacterial immune evasion.
- This approach facilitates the study of infection biology and the identification of therapeutic targets.
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