Subject(s)
Genomics/methods , Microscopy, Atomic Force/methods , Animals , Biomarkers/analysis , Genetic Predisposition to Disease , Genetic Variation , Genomics/economics , High-Throughput Nucleotide Sequencing/economics , High-Throughput Nucleotide Sequencing/methods , Humans , Microscopy, Atomic Force/economicsABSTRACT
Fast, high-resolution mapping of heterogeneous interfaces with a wide elastic modulus range is a major goal of atomic force microscopy (AFM). This goal becomes more challenging when the nanomechanical mapping involves biomolecules in their native environment. Over the years, several AFM-based methods have been developed to address this goal. However, none of these methods combine sub-nanometer spatial resolution, quantitative accuracy, fast data acquisition speed, wide elastic modulus range and operation in physiological solutions. Here, we present detailed procedures for generating high-resolution maps of the elastic properties of biomolecules and polymers using bimodal AFM. This requires the simultaneous excitation of the first two eigenmodes of the cantilever. An amplitude modulation (AM) feedback acting on the first mode controls the tip-sample distance, and a frequency modulation (FM) feedback acts on the second mode. The method is fast because the elastic modulus, deformation and topography images are obtained simultaneously. The method is efficient because only a single data point per pixel is needed to generate the aforementioned images. The main stages of the bimodal imaging are sample preparation, calibration of the instrument, tuning of the microscope and generation of the nanomechanical maps. In addition, with knowledge of the deformation, bimodal AFM enables reconstruction of the true topography of the surface. It takes ~9 h to complete the whole procedure.
Subject(s)
Elasticity Imaging Techniques/methods , Elasticity , Microscopy, Atomic Force/methods , Polymers/chemistry , Proteins/chemistry , Animals , Biocompatible Materials/chemistry , Biomechanical Phenomena , Elasticity Imaging Techniques/economics , Elasticity Imaging Techniques/instrumentation , Equipment Design , Halobacterium salinarum/chemistry , Halobacterium salinarum/ultrastructure , Humans , Microscopy, Atomic Force/economics , Microscopy, Atomic Force/instrumentation , Models, Molecular , Proteasome Endopeptidase Complex/chemistry , Proteasome Endopeptidase Complex/ultrastructure , Proteins/ultrastructure , Purple Membrane/chemistry , Purple Membrane/ultrastructure , Time FactorsABSTRACT
A rapid and facile signal enhancement method for detecting alpha-fetoprotein (AFP) was developed using the magnetic agglomeration of ferromagnetic nanoparticles and microcantilever sensors. The resonance frequency and deflection of the cantilevers were found to be more than 10-fold greater than that before physical agglomeration of the free nanoparticles around the magnetized nanoparticles.
Subject(s)
Immunoassay/instrumentation , Magnets/chemistry , Microscopy, Atomic Force/instrumentation , Nanoparticles/chemistry , alpha-Fetoproteins/analysis , Antibodies, Immobilized/immunology , Biosensing Techniques/economics , Biosensing Techniques/instrumentation , Humans , Immunoassay/economics , Microscopy, Atomic Force/economics , Sensitivity and Specificity , Time Factors , alpha-Fetoproteins/immunologyABSTRACT
Atomic force microscopy (AFM) can be used to co-localize mechanical properties and topographical features through property mapping techniques. The most common approach for testing biological materials at the microscale and nanoscale is force mapping, which involves taking individual force curves at discrete sites across a region of interest. The limitations of force mapping include long testing times and low resolution. While newer AFM methodologies, like modulated scanning and torsional oscillation, circumvent this problem, their adoption for biological materials has been limited. This could be due to their need for specialized software algorithms and/or hardware. The objective of this study is to develop a novel force scanning technique using AFM to rapidly capture high-resolution topographical images of soft biological materials while simultaneously quantifying their mechanical properties. Force scanning is a straightforward methodology applicable to a wide range of materials and testing environments, requiring no special modification to standard AFMs. Essentially, if a contact-mode image can be acquired, then force scanning can be used to produce a spatial modulus map. The current study first validates this technique using agarose gels, comparing results to ones achieved by the standard force mapping approach. Biologically relevant demonstrations are then presented for high-resolution modulus mapping of individual cells, cell-cell interfaces, and articular cartilage tissue.
