Development of Binding Assays in Microfabricated Picoliter Vials:  An Assay for Biotin

Grosvenor, Anne L.; Feltus, Agatha; Conover, Richard C.; Daunert, Sylvia; Anderson, Kimberly W.

doi:10.1021/ac991289+

Cited by 22 publications

(19 citation statements)

References 29 publications

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“…Results of the dose-response curve indicated that decreasing concentrations of cortisol, and corresponding increasing concentrations of free antibody, resulted in an increase in the bioluminescent signal from the conjugate (Figure 4). One may initially assume that increasing the amount of antibody available to bind the conjugate would result in sterically hindering aequorin and, thereby, reduce its bioluminescence, as shown in the homogeneous aequorin-based assay for biotin (21). However, a similar homogeneous response to ours, wherein signal increased with increasing antibody concentration, was found by Lewis and colleagues when they studied the response of an aequorin-thyroxine conjugate to increasing concentrations of antithyroxine antibody (22).…”

Section: Discussionsupporting

confidence: 61%

Aequorin-Based Homogeneous Cortisol Immunoassay for Analysis of Saliva Samples

et al. 2007

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Homogeneous assays are attractive because they are performed in only one phase, namely, the liquid phase, and thus, they do not require separation of phases as their heterogeneous counterparts do. As opposed to heterogeneous assays, the signal generation in a homogeneous assay is a direct result of analyte binding, which allows the multiple washing and incubation steps required in an indirect heterogeneous assay format to be eliminated. Moreover, homogeneous assays are usually fast and amenable to miniaturization and automation. In this article, we describe the development of a homogeneous assay for the hormone cortisol using the bioluminescent photoprotein aequorin as a reporter molecule. A cortisol derivative was chemically conjugated to the lysine residues of a genetically modified aequorin in order to prepare an aequorin-cortisol conjugate capable of binding anticortisol antibodies. The binding of anticortisol antibodies to the aequorin-cortisol conjugate resulted in a linear response reflected in the emission of bioluminescence by aequorin. A competitive binding assay was developed by simultaneously incubating the aequorin-cortisol conjugate, the anticortisol antibodies, and the sample containing free cortisol. Dose-response curves were generated relating the intensity of the bioluminescence signal with the concentration of free cortisol in the sample. The optimized homogeneous immunoassay produced a detection limit of 1 x 10 (-10) M of free cortisol, with a linear dynamic range spanning from 1 x 10 (-5) to 1 x 10 (-9) M. Both serum and salivary levels of cortisol fall well within this assay's linear range (3.0 x 10 (-7) M to 7.5 x 10 (-7) M and 1.0 x 10 (-8) M to 2.5 x 10 (-8) M, respectively), thereby making this assay attractive for the analysis of this hormone in biological samples. To that end, it was demonstrated that the assay can be reliably used to measure the concentration of free cortisol in saliva without significant pretreatment of the sample.

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Section: Discussionsupporting

confidence: 61%

Aequorin-Based Homogeneous Cortisol Immunoassay for Analysis of Saliva Samples

et al. 2007

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“…The integration of such structures with fluids and biological materials is enabling multiple applications. For example, small volume reaction containers are being considered for high‐throughput screening, (Grosvenor et al , 2000; Angenendt et al , 2005), single molecule enzymology (Rondelez et al , 2005; Rissin and Walt, 2006), and analyses of single cells (Cooper, 1999; Johannessen et al , 2002). These reaction containers are also being developed for the cell‐free synthesis of proteins (Nojima et al , 2000; Tabuchi et al , 2002; Yamamoto et al , 2002; Angenendt et al , 2004; Kinpara et al , 2004; Mei et al , 2005).…”

Section: Nanomaterials: From Individual Elements To Cell‐like Complexitymentioning

confidence: 99%

Nano‐enabled synthetic biology

Doktycz

Simpson

2007

Molecular Systems Biology

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Biological systems display a functional diversity, density and efficiency that make them a paradigm for synthetic systems. In natural systems, the cell is the elemental unit and efforts to emulate cells, their components, and organization have relied primarily on the use of bioorganic materials. Impressive advances have been made towards assembling simple genetic systems within cellular scale containers. These biological system assembly efforts are particularly instructive, as we gain command over the directed synthesis and assembly of synthetic nanoscale structures. Advances in nanoscale fabrication, assembly, and characterization are providing the tools and materials for characterizing and emulating the smallest scale features of biology. Further, they are revealing unique physical properties that emerge at the nanoscale. Realizing these properties in useful ways will require attention to the assembly of these nanoscale components. Attention to systems biology principles can lead to the practical development of nanoscale technologies with possible realization of synthetic systems with cell-like complexity. In turn, useful tools for interpreting biological complexity and for interfacing to biological processes will result.

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“…Self-contained micropumps are preferred for manipulating a minute amount of fluid with high precision in a microfluidic systems [9]. Lab-on-a-chip applications often require the manipulation control of of fluid volume on the order of 1 pL [10][11][12]. To avoid dead volume, a micropump should be integrated on the same chip with other components.…”

Section: Introductionmentioning

confidence: 99%

Liquid metal microcoils for sensing and actuation in lab-on-a-chip applications

Kong

Nguyen

2013

Microsyst Technol

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The use of metals and alloys with melting point near room temperature, called here as liquid metals, allows the integration of complex three-dimensional metallic micro structures in lab-on-chip devices. The process involves the injection molten liquid metal into microchannels and subsequent solidification at room temperature. The paper reports a technique for the fabrication of threedimensional multilayer liquid-metal microcoils by lamination of dry adhesive films. The adhesive-based liquid metal microcoil could be used for magnetic resonance relaxometry (MRR) measurement in a lab-on-a-chip platform. Not only that the coil has a low direct-current resistance, it also has a high quality factor. In this paper, we investigate the sensing and actuating capabilities of the liquid metal microcoil. The sensing capability of the microcoil is demonstrated with the coil working as a blood hematocrit level sensor. In a magnetic resonance relaxometry measurement, the transverse relaxation rate of the blood sample increases quadratically with the hematocrit level due to higher magnetic susceptibility. Furthermore, a vibrating adhesive membrane with the embedded coil was realized for electromagnetic actuation. A maximum deflection of approximately 50 microns at a low resonance frequency of 15 Hz can be achieved with a maximum driving current of 300 mA.

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Development of Binding Assays in Microfabricated Picoliter Vials: An Assay for Biotin

Cited by 22 publications

References 29 publications

Aequorin-Based Homogeneous Cortisol Immunoassay for Analysis of Saliva Samples

Aequorin-Based Homogeneous Cortisol Immunoassay for Analysis of Saliva Samples

Nano‐enabled synthetic biology

Liquid metal microcoils for sensing and actuation in lab-on-a-chip applications

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