An Advanced Biosensor for Molecular Interaction Studies

用于分子相互作用研究的先进生物传感器

• 批准号: 7999170
• 负责人: DARRYL J. BORNHOP
• 金额: $ 25.69万
• 依托单位: MOLECULAR SENSING, INC.
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-09-01 至 2011-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7999170
• 关键词: Address; Adoption; Affinity; Algorithms; Antibodies; Antigens; Area; Back; Binding; Biochemical; Biological Assay; Biological Markers; Biological Sciences; Biology; Biomedical Research; Biosensor; Bread; Calmodulin; Calorimetry; Catalysis; Cell physiology; Cells; Chemicals; Chemistry; Communities; Complex; Computer software; Consumption; Data; Data Collection; Data Quality; Detection; Development; Diagnosis; Diagnostic; Dimensions; Dimethyl Sulfoxide; Drug Delivery Systems; Environment; Fees; Financial compensation; Foundations; Fourier Transform; Future; G Protein-Coupled Receptor Genes; Generations; Glass; Glycerol; Grant; Hand functions; Human; Immobilization; In Vitro; Interferometry; Kinetics; Knowledge; Label; Laboratories; Lasers; Lead; Liquid substance; Longevity; Measurement; Measures; Methodology; Methods; Microfluidics; Modeling; Molecular; Monitor; Noise; Optics; Orphan; Pattern; Performance; Pharmaceutical Preparations; Pharmacologic Substance; Phase; Plasma; Plastics; Positioning Attribute; Proteins; Radioactive; Reaction; Refractive Indices; Relative (related person); Research; Research Contracts; Research Personnel; Sampling; San Francisco; Scheme; Screening procedure; Serum; Signal Transduction; Silicon Dioxide; Solutions; Specimen; Staging; Surface; System; Techniques; Technology; Technology Transfer; Temperature; Therapeutic; Thermodynamics; Titrations; Training; base; biological research; commercialization; cost; drug discovery; experience; extracellular; graphical user interface; improved; inhibitor/antagonist; insight; instrument; instrumentation; miniaturize; novel; programs; prototype; public health relevance; receptor; sensor; small molecule; tool; user-friendly

DESCRIPTION (provided by applicant): Molecular interactions are central to the understanding of basic biology and cellular function, are paramount to all molecular assays (research and in vitro diagnostics), are essential in biomarker discovery and critical toward evaluating therapeutics. However, the tools available to quantify these interactions have limitations, in so far as they either rely upon altering the native state of the interacting molecules by incorporating labeling schemes, demanding surface immobilization of one of the interacting moieties or require prohibitive quantities of sample. The ability to perform pure liquid-phase molecular binding analysis at high sensitivity, in a miniaturized format, is therefore desirable and has now been demonstrated with back-scattering interferometry (BSI). BSI employs a simple and inexpensive optical train comprised of a He-Ne laser, a microfluidic channel, and a position sensor monitoring minute refractive index changes. These measurements may be made within a microfluidic channel formed in glass, fused silica, or plastic and at physiologically relevant concentrations in complex matrices. Yet the current embodiment of BSI is limited. Tedious alignment methods, immature transduction schemes, poorly refined sample handling and introduction methods, expensive disposable chips, and performance limitations due to environmental noise sensitivity, all impede the wide dissemination and adoption of BSI in the life science and drug discovery communities. We propose to refine BSI, facilitating commercialization by Molecular Sensing Inc. and allowing the subsequent broad dissemination in the biological research community (under a future Phase II grant). Under this Phase I project, we will enable transfer to the industrial sector by performing three specific aims: 1) constructing an optically simple two channel BSI for enhanced S/N and environmental noise compensation, 2) simplifying alignment and improving performance through the use of a cross-correlation based position sensing algorithm and 3) implementing a user friendly graphical interface (GUI). Model interaction systems will be used to demonstrate that BSI gives meaningful and quantitative binding affinity values (from <M to pM) and that it can be used to screen for molecular interactions in various matrices (serum, cell-free media, DMSO). PUBLIC HEALTH RELEVANCE: Proteins interact constantly with one another and with other entities in the cellular and extracellular environment. However, sensitive in vitro methods to monitor such interactions, in particular methods that are label-free and do not involve surface immobilization, have until recently been lacking. Free-solution, label-free, molecular interactions can now be investigated using back-scattering interferometry (BSI), allowing quantification of KD values ranging from micromolar to picomolar using miniscule quantities of the binding pairs. With BSI, little a priori knowledge about the proteins, antibodies, or drug targets etc. is necessary to quantify the level of interaction and screen for efficacy or perform a diagnosis. The BSI methodology has the potential to shift the paradigm for molecular interaction studies, allowing screening for unknown binding partners of orphan receptors, study of inhibitors for GPCR targets, as well as for numerous other binding pairs and is likely applicable to diagnostics and therapeutic monitoring in human specimens. Furthermore, the technology is simple and inexpensive allowing for the potential of BSI to be widely disseminated.

描述（由申请人提供）：分子相互作用对于理解基础生物学和细胞功能至关重要，对于所有分子测定（研究和体外诊断）至关重要，对于生物标志物发现至关重要，对于评价治疗至关重要。然而，可用于量化这些相互作用的工具具有局限性，因为它们要么依赖于通过掺入标记方案来改变相互作用分子的天然状态，要求相互作用部分之一的表面固定化，要么需要禁止量的样品。因此，在高灵敏度下以小型化形式进行纯液相分子结合分析的能力是期望的，并且现在已经用背散射干涉法（BSI）证明。BSI采用了一种简单而廉价的光学系统，包括He-Ne激光器、微流体通道和监测微小折射率变化的位置传感器。这些测量可以在玻璃、熔融石英或塑料中形成的微流体通道内进行，并且在复杂基质中以生理相关浓度进行。然而，BSI的当前实施例是有限的。繁琐的比对方法、不成熟的转导方案、不完善的样品处理和引入方法、昂贵的一次性芯片以及由于环境噪声敏感性而导致的性能限制，都阻碍了BSI在生命科学和药物发现领域的广泛传播和采用。我们建议完善BSI，促进Molecular Sensing Inc.的商业化。并允许随后在生物研究界广泛传播（根据未来的第二阶段赠款）。在这个第一阶段项目中，我们将通过执行三个具体目标实现向工业部门的转移：1）构建光学简单的双通道BSI，以增强S/N和环境噪声补偿，2）通过使用基于互相关的位置传感算法简化对准并提高性能，以及3）实现用户友好的图形界面（GUI）。模型相互作用系统将用于证明BSI给出有意义的定量结合亲和力值（从<M到pM），并且其可用于筛选各种基质（血清、无细胞培养基、DMSO）中的分子相互作用。 公共卫生相关性：蛋白质相互之间以及与细胞和细胞外环境中的其他实体不断相互作用。然而，敏感的体外方法来监测这种相互作用，特别是方法，是无标记的，不涉及表面固定，直到最近一直缺乏。自由溶液，无标记，分子相互作用，现在可以使用背散射干涉法（BSI）进行研究，允许定量的KD值范围从微摩尔到皮摩尔使用微量的结合对。对于BSI，几乎不需要关于蛋白质、抗体或药物靶点等的先验知识来量化相互作用的水平并筛选疗效或进行诊断。BSI方法有可能改变分子相互作用研究的范式，允许筛选孤儿受体的未知结合伴侣，研究GPCR靶点的抑制剂以及许多其他结合对，并可能适用于人体标本的诊断和治疗监测。此外，该技术简单而廉价，使BSI的潜力得到广泛传播。