Multiplexed Nucleic Acid programmable protein arrays

Xiaobo Yu, Lusheng Song, Brianne Petritis, Xiaofang Bian, Haoyu Wang, Jennifer Viloria, Jin Park, Hoang Bui, Han Li, Jie Wang, Lei Liu, Liuhui Yang, Hu Duan, David N. McMurray, Jacqueline M. Achkar, Mitch Magee, Ji Qiu, Joshua LaBaer

Research output: Contribution to journalArticlepeer-review

19 Scopus citations


Rationale: Cell-free protein microarrays display naturally-folded proteins based on just-in-time in situ synthesis, and have made important contributions to basic and translational research. However, the risk of spot-to-spot cross-talk from protein diffusion during expression has limited the feature density of these arrays. Methods: In this work, we developed the Multiplexed Nucleic Acid Programmable Protein Array (M-NAPPA), which significantly increases the number of displayed proteins by multiplexing as many as five different gene plasmids within a printed spot. Results: Even when proteins of different sizes were displayed within the same feature, they were readily detected using protein-specific antibodies. Protein-protein interactions and serological antibody assays using human viral proteome microarrays demonstrated that comparable hits were detected by M-NAPPA and non-multiplexed NAPPA arrays. An ultra-high density proteome microarray displaying > 16k proteins on a single microscope slide was produced by combining M-NAPPA with a photolithography-based silicon nano-well platform. Finally, four new tuberculosis-related antigens in guinea pigs vaccinated with Bacillus Calmette-Guerin (BCG) were identified with M-NAPPA and validated with ELISA. Conclusion: All data demonstrate that multiplexing features on a protein microarray offer a cost-effective fabrication approach and have the potential to facilitate high throughput translational research.

Original languageEnglish (US)
Article number20151
Issue number16
StatePublished - 2017


  • Antibody
  • Biomarker
  • Cell-free protein microarray
  • Protein-protein interaction
  • Proteomics

ASJC Scopus subject areas

  • Medicine (miscellaneous)
  • Pharmacology, Toxicology and Pharmaceutics (miscellaneous)


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