Whole-Cell Sensing for a Harmful Bloom-Forming Microscopic Alga by Measuring Antibody–Antigen Forces

Lee, A.S.; Mahapatro, Mrinal; Caron, David A.; Requicha, Ari; Stauffer, Beth; Thompson, Mark E.; Zhou, Chongwu

doi:10.1109/tnb.2006.880767

Cited by 8 publications

(8 citation statements)

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“…Pioneer studies have been performed, especially on more stable interactions such as biotin−avidin, − ,,,, antigens−antibodies, ,,, or receptors−ligands. , Afterward, also many other kinds of biological complexes, different for stability and composition, such as DNA−proteins, − transient complexes, , chaperonins with their substrates, or the exotic p53−azurin, have been successfully explored by this technique. Some recent works show the possibility of investigating the interaction between ligands and membrane receptors, probing directly the whole living cells. − This approach, allowing for mapping of the presence and location of specific receptors on the membrane, provides details about the cellular functional state.…”

Section: Molecular Recognition and Dynamic Force Spectroscopymentioning

confidence: 99%

Atomic Force Spectroscopy in Biological Complex Formation: Strategies and Perspectives

Bizzarri

Cannistraro

2009

J. Phys. Chem. B

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Atomic force spectroscopy has become a widely used technique for investigating forces, energies, and dynamics of biomolecular interactions. These studies provide dissociation kinetic parameters by pulling apart proteins involved in a complex. Biological complexes are studied under near-physiological conditions, without labeling procedures, and are probed one at time, the latter allowing to one obtain results which are not averaged over the ensemble. However, to gain reliable information, some experimental aspects have to be carefully controlled. In particular, the immobilization of molecular partners to AFM tips and supports, required to force the molecular dissociation, plays a crucial role in determining the success of the experiments. To actually resolve single interactions, multiple simultaneous complex dissociations have to be avoided, and nonspecific adhesions, commonly found in these studies, have to be recognized and discarded. This article is aimed at offering a critical revisitation of the atomic force spectroscopy technique applied to the study of biomolecular interactions, highlighting the critical points, identifying strategies to be adopted for a more reliable data extraction and interpretation, and pointing out the experimental and theoretical aspects which still need to be refined. To this purpose, we take advantage of the vast landscape of literature and then proceed into the details of our works. In this respect, we describe the general principles of the technique, the procedures for protein immobilization, and how they can affect the results. We emphasize the use of computational docking to predict molecular complex configurations, when unknown, as a useful approach to select proper anchorage architectures. Additionally, we deal with data acquirement and analysis, with regard to the force curve selection, to the force histograms interpretation, and to the theoretical frameworks used to extract kinetic parameters. Through this, we outline that AFS can be successfully used both to investigate complexes having very different affinities and also to reveal competitive binding mechanisms, thus gaining deeper information about molecular interactions.

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Section: Molecular Recognition and Dynamic Force Spectroscopymentioning

confidence: 99%

Atomic Force Spectroscopy in Biological Complex Formation: Strategies and Perspectives

Bizzarri

Cannistraro

2009

J. Phys. Chem. B

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“…Therefore, we examined the effect of directly diluting seawater samples in PBS-T as an alternative to filtration/centrifugation. A 1:10 10,20,30,45, or 60 min showed little variation of staining intensity between 10 and 30 min. The mean FL1 signal increased slightly with prolonged incubation times (40 to 60 min), but the increase was not significant (bootstrap-t analysis, P Ͼ 0.05), and stained cells were easily distinguishable on flow cytograms from unstained cells, even with shorter incubation times ( Table 1).…”

Section: Resultsmentioning

confidence: 89%

“…These approaches have included immunology-based (3,7,30) and genetics-based detection protocols (41), and each method has inherent advantages, disadvantages, and appropriate ecological applications. The immunofluorescence microscopy method of Anderson et al (3) provided the first method to accurately identify and count A. anophagefferens organisms in natural algal assemblages through the use of immunofluorescence microscopy.…”

Section: Discussionmentioning

confidence: 99%

“…The ELISA technique has a high rate of sample throughput relative to the polyclonal microscopy method but also has a somewhat higher limit of detection (approximately 5,000 cells ml Ϫ1 ). A recent application of the same monoclonal antibody enabled single-cell detection by atomic force microscopy (30). This approach demonstrates the potential to build sensors with single-cell resolution, but its application to natural water samples is not yet technically feasible.…”

Section: Discussionmentioning

confidence: 99%

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Immunofluorescence Flow Cytometry Technique for Enumeration of the Brown-Tide Alga, Aureococcus anophagefferens

Stauffer

Schaffner

Wazniak

et al. 2008

Appl Environ Microbiol

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A new immunologically based flow cytometry (IFCM) technique was developed to enumerate Aureococcus anophagefferens, a small pelagophyte alga that is the cause of "brown tides" in bays and estuaries of the mid-Atlantic states along the U.S. coast. The method utilizes a monoclonal antibody conjugated to fluorescein isothiocyanate (FITC-MAb) to label the surface of A. anophagefferens cells which are then detected and enumerated by using a flow cytometer. Optimal conditions for FITC-MAb staining, including solution composition, incubation times, and FITC-MAb concentrations, were determined. The FITC-MAb method was tested for cross-reactivity with nontarget, similarly sized, photoautotrophic protists, and the method was compared to an enzyme-linked immunosorbent assay (ELISA) using the same MAb. Comparisons of the IFCM technique to traditional microscopy enumeration of cultures and spiked environmental samples showed consistent agreement over several orders of magnitude (r 2 > 0.99). Comparisons of the IFCM and ELISA techniques for enumerating cells from a predation experiment showed a substantial overestimation (up to 10 times higher) of the ELISA in the presence of consumers of A. anophagefferens, presumably due to egested cell fragments that retained antigenicity, using the ELISA method, but were not characterized as whole algal cells by the IFCM method. Application of the IFCM method to environmental "brown-tide" samples taken from the coastal bays of Maryland demonstrated its efficacy in resolving A. anophagefferens abundance levels throughout the course of a bloom and over a large range of abundance values. IFCM counts of the brown-tide alga from natural samples were consistently lower than those obtained using the ELISA method and were equivalent to those of the polyclonal immunofluorescence microscopy technique, since both methods discriminate intact cells. Overall, the IFCM approach was an accurate and relatively simple technique for the rapid enumeration of A. anophagefferens in natural samples over a wide range of abundance values (10 3 to 10 6 cells ml ؊1 ).

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“…Recent developments in biomolecular computing have demonstrated the feasibility of biocomputers [33], a promising first step toward future nanoprocessors. Other examples include studies of building biosensors and nanokinetic devices [7,8,13,19,34].…”

Section: Introductionmentioning

confidence: 99%

Nanorobot architecture for medical target identification

Cavalcanti¹,

Shirinzadeh²,

Freitas³

et al. 2007

Nanotechnology

166

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This work has an innovative approach for the development of nanorobots with sensors for medicine. The nanorobots operate in a virtual environment comparing random, thermal and chemical control techniques. The nanorobot architecture model has nanobioelectronics as the basis for manufacturing integrated system devices with embedded nanobiosensors and actuators, which facilitates its application for medical target identification and drug delivery. The nanorobot interaction with the described workspace shows how time actuation is improved based on sensor capabilities. Therefore, our work addresses the control and the architecture design for developing practical molecular machines. Advances in nanotechnology are enabling manufacturing nanosensors and actuators through nanobioelectronics and biologically inspired devices. Analysis of integrated system modeling is one important aspect for supporting nanotechnology in the fast development towards one of the most challenging new fields of science: molecular machines. The use of 3D simulation can provide interactive tools for addressing nanorobot choices on sensing, hardware architecture design, manufacturing approaches, and control methodology investigation.

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Whole-Cell Sensing for a Harmful Bloom-Forming Microscopic Alga by Measuring Antibody–Antigen Forces

Cited by 8 publications

References 50 publications

Atomic Force Spectroscopy in Biological Complex Formation: Strategies and Perspectives

Atomic Force Spectroscopy in Biological Complex Formation: Strategies and Perspectives

Immunofluorescence Flow Cytometry Technique for Enumeration of the Brown-Tide Alga, Aureococcus anophagefferens

Nanorobot architecture for medical target identification

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