Live Cell Interferometry Reveals Cellular Dynamism During Force Propagation

Department of Chemistry and Biochemistry, University of California, Los Angeles, 607 Charles Young Drive East, Los Angeles, California 90095, USA.
ACS Nano (Impact Factor: 12.88). 06/2008; 2(5):841-6. DOI: 10.1021/nn700303f
Source: PubMed


Cancer and many other diseases are characterized by changes in cell morphology, motion, and mechanical rigidity. However, in live cell cytology, stimulus-induced morphologic changes typically take 10-30 min to detect. Here, we employ live-cell interferometry (LCI) to visualize the rapid response of a whole cell to mechanical stimulation, on a time scale of seconds, and we detect cytoskeletal remodeling behavior within 200 s. This behavior involved small, rapid changes in cell content and miniscule changes in shape; it would be difficult to detect with conventional or phase contrast microscopy alone and is beyond the dynamic capability of AFM. We demonstrate that LCI provides a rapid, quantitative reconstruction of the cell body with no labeling. This is an advantage over traditional microscopy and flow cytometry, which require cell surface tagging and/or destructive cell fixation for labeling.

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    • "Live cell interferometry (LCI) is a label-free optical microscopy technique which measures whole cell responses. LCI uses a Michelson-type interferometer to compare the optical thickness of living cells in a sample chamber to the optical thickness of fluid in a reference chamber in order to quantify the optical thickness difference between a cell and its surrounding media [12], [13]. The optical thickness difference due to the interaction of light with cellular biomass is linearly proportional to the material density of a cell [14]. "
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    ABSTRACT: Existing approaches that quantify cytotoxic T cell responses rely on bulk or surrogate measurements which impede the direct identification of single activated T cells of interest. Single cell microscopy or flow cytometry methodologies typically rely on fluorescent labeling, which limits applicability to primary cells such as human derived T lymphocytes. Here, we introduce a quantitative method to track single T lymphocyte mediated cytotoxic events within a mixed population of cells using live cell interferometry (LCI), a label-free microscopy technique that maintains cell viability. LCI quantifies the mass distribution within individual cells by measuring the phase shift caused by the interaction of light with intracellular biomass. Using LCI, we imaged cytotoxic T cells killing cognate target cells. In addition to a characteristic target cell mass decrease of 20-60% over 1-4 h following attack by a T cell, there was a significant 4-fold increase in T cell mass accumulation rate at the start of the cytotoxic event and a 2-3 fold increase in T cell mass relative to the mass of unresponsive T cells. Direct, label-free measurement of CD8+ T and target cell mass changes provides a kinetic, quantitative assessment of T cell activation and a relatively rapid approach to identify specific, activated patient-derived T cells for applications in cancer immunotherapy.
    PLoS ONE 07/2013; 8(7):e68916. DOI:10.1371/journal.pone.0068916 · 3.23 Impact Factor
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    • "The prolonged poking of every single cell can result in the remodeling of the cytoskeleton (Reed et al., 2008). The influence of the prolonged poking on a single cell can be easily checked by plotting the dependence between the obtained Young's modulus and the time elapsed during the indentation experiment. "
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    ABSTRACT: The major characteristics of cancer metastasis is the ability of the primary tumor cells to migrate by way of the blood or lymph vessels and to form tumors at multiple, distant sites. There are evidences that cancer progression is characterized by disruption and/or reorganization of cytoskeleton (i.e. cellular scaffold). This is accompanied by various molecular alterations influencing the overall mechanical resistance of cells. Current approach in diagnosis focuses mainly on microbiological, immunological, and pathological aspects rather than on the biomechanics of diseases. The determination of mechanical properties of an individual living cell has became possible with the development of local measurement techniques, such as atomic force microscopy, magnetic or optical tweezers. The advantage of them lies in the capability to measure living cells at a single cell level and in liquid conditions, close to natural environment.
    Micron 02/2012; 43(12):1259-66. DOI:10.1016/j.micron.2012.01.019 · 1.99 Impact Factor
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    • "The current generation of AFMs can be integrated with complementary methodologies, including ionic conductance [87], total internal reflection fluorescence (TIRF) [88,89], fluorescence resonance energy transfer (FRET) [90] and other physico-chemical measurements [91], thereby enabling detailed structure-function studies of biofilms, dental surfaces and implants. Rapid quantitative changes in oral surfaces and cell morphology, motion and mechanical rigidity via live-cell interferometry (LCI) [92], can also be combined with the dynamic capability of AFM. Depending on the specific interests and technical requirements, the variety of combination techniques available with AFM would cover both transparent (e.g., dental biofilms) and non-transparent samples such as dental implants and fillings. "
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    ABSTRACT: About 80% of US adults have some form of dental disease. There are a variety of new dental products available, ranging from implants to oral hygiene products that rely on nanoscale properties. Here, the application of AFM (Atomic Force Microscopy) and optical interferometry to a range of dentistry issues, including characterization of dental enamel, oral bacteria, biofilms and the role of surface proteins in biochemical and nanomechanical properties of bacterial adhesins, is reviewed. We also include studies of new products blocking dentine tubules to alleviate hypersensitivity; antimicrobial effects of mouthwash and characterizing nanoparticle coated dental implants. An outlook on future "nanodentistry" developments such as saliva exosomes based diagnostics, designing biocompatible, antimicrobial dental implants and personalized dental healthcare is presented.
    International Journal of Molecular Sciences 06/2010; 11(6):2523-45. DOI:10.3390/ijms11062523 · 2.86 Impact Factor
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