High-Speed Lens-Free Holographic Sensing of Protein Molecules Using Quantitative Agglutination Assays

Zhen Xiong, Colin J. Potter, Euan McLeod

Research output: Contribution to journalArticlepeer-review

10 Scopus citations


Accurate, cost-effective, easy-to-use, and point-of-care sensors for protein biomarker levels are important for disease diagnostics. A cost-effective and compact readout approach that has been used for several diagnostic applications is lens-free holographic microscopy, which provides an ultralarge field of view and submicron resolution when it is coupled with pixel super-resolution techniques. Despite its potential as a diagnostic technique, lens-free microscopy has not previously been applied to quantitative protein molecule sensing in solution, which can simplify sensing protocols and ultimately enable measurements of binding kinetics in physiological conditions. Here, we sense interferon-γ(an immune system biomarker) and NeutrAvidin molecules in solution by combining lens-free microscopy with a one-step bead-based agglutination assay, enabled by a custom high-speed light-emitting diode (LED) array and automated image processing routines. We call this a quantitative large-area binding (QLAB) sensor. The high-speed light source provides, for the first time, pixel super-resolved imaging of >104 2 μm beads in solution undergoing Brownian motion, without significant motion blur. The automated image processing routines enable the counting of individual beads and clusters, providing a quantitative sensor readout that depends on both bead and analyte concentrations. Fits to the chemical binding theory are provided. For NeutrAvidin, we find a limit of detection (LOD) of <27 ng/mL (450 pM) and a dynamic range of 2-4 orders of magnitude. For mouse interferon-γ, the LOD is <3 ng/mL (200 pM) and the dynamic range is at least 4 orders of magnitude. The QLAB sensor holds promise for point-of-care applications in low-resource communities and where protocol simplicity is important.

Original languageEnglish (US)
Pages (from-to)1208-1217
Number of pages10
JournalACS Sensors
Issue number3
StatePublished - Mar 26 2021


  • Brownian motion
  • agglutination assay
  • high-speed super-resolution
  • lens-free holographic microscopy
  • point-of-care
  • protein biomarkers

ASJC Scopus subject areas

  • Bioengineering
  • Instrumentation
  • Process Chemistry and Technology
  • Fluid Flow and Transfer Processes


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