Wei Shan Tan, Arthur M. de Jong, Menno W. J. Prins
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Revealing Spatial Molecular Heterogeneity of High-Density Biofunctionalized Surfaces Using DNA-PAINT
The quantification and control of molecular densities and distributions on biofunctionalized surfaces are key for enabling reproducible functions in biosciences. Here, we describe an analysis methodology for quantifying the density and spatial distribution of high-density biofunctionalized surfaces, with densities in the order of 102–105 biomolecules per μm2 area, in a short measurement time. The methodology is based on single-molecule DNA-PAINT imaging combined with simulation models that compensate for lifetime and spatial undersampling effects, resulting in three distinct molecule counting methods and a statistical test for spatial distribution. The analysis methodology is exemplified for a surface with ssDNA affinity binder molecules coupled to a PLL-g-PEG antifouling coating. The results provide insights into the biofunctionalization efficiency, yield, and homogeneity. Furthermore, the data reveal that heterogeneity is inherent to the biofunctionalization process and shed light on the underlying molecular mechanisms. We envision that DNA-PAINT imaging with the developed analysis framework will become a versatile tool to study spatial heterogeneity of densely biofunctionalized surfaces for a wide range of applications.
期刊介绍:
ACS Applied Materials & Interfaces is a leading interdisciplinary journal that brings together chemists, engineers, physicists, and biologists to explore the development and utilization of newly-discovered materials and interfacial processes for specific applications. Our journal has experienced remarkable growth since its establishment in 2009, both in terms of the number of articles published and the impact of the research showcased. We are proud to foster a truly global community, with the majority of published articles originating from outside the United States, reflecting the rapid growth of applied research worldwide.