Tunable affinity and heat stable antibody targeting of nanoparticles

纳米颗粒的可调亲和力和热稳定性抗体靶向

• 批准号: 8096287
• 负责人: Todd Sulchek
• 金额: $ 22.75万
• 依托单位: GEORGIA INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-04-01 至 2013-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8096287
• 关键词: Address; Adhesions; Affinity; Agglutination; Antibodies; Antibody Affinity; Antigen Targeting; Antigens; Architecture; Atomic Force Microscopy; Bacillus anthracis; Bacillus anthracis spore; Binding; Binding Sites; Biological Assay; Biological Warfare; Capsid Proteins; Detection; Development; Diagnosis; Dissociation; Drug Delivery Systems; Effectiveness; Engineering; Fc Receptor; Flow Cytometry; Goals; Heating; High temperature of physical object; Image; Immunoglobulin G; Kinetics; Length; Lymphocyte; Measurement; Measures; Methods; Modification; Molecular; Organism; Particle Size; Pharmacologic Substance; Polymers; Property; Reagent; Recombinants; Research Proposals; Scheme; Specificity; Spectrum Analysis; Surface Plasmon Resonance; Techniques; Temperature; Vertebrates; antigen binding; base; design; detector; improved; microbial; microorganism; nano; nanoparticle; nanoscale; particle; pathogen; physical property; receptor; sensor; single molecule

DESCRIPTION (provided by applicant): Our proposed study will demonstrate that we can produce a targeting vehicle capable of both tunable affinity and heat-stability through nanoscale engineering of Variable Lymphocyte Receptors (VLR) antibodies. We will demonstrate these targeting qualities through binding assays of VLR functionalized nano/microparticles. We will create multivalent VLR antibodies with a range of valencies, linker lengths, and that have suitable residues for attachment to nano/microparticles. The multivalent recombinant forms of VLRs will recognize the Bacillus anthracis spore coat protein called BclA. We will verify the proper valency and linker arrangement with biophysical techniques and single molecule imaging. We will measure the binding strength the VLR antibodies to the BclA antigen with surface plasmon resonance (SPR) and single-molecule binding measurements based upon atomic force microscopy force spectroscopy. We will measure the kinetic rates and the resulting affinity of the multivalent antibodies as a function of valency and linker length. We will also validate that antigen binding occurs even when raised to high temperatures. We will conjugate the multivalent VLRs to several micro/nanoparticles and validate adhesion to B. anthracis using high throughput flow cytometry. We will measure particle adhesion and agglutination as a function of a) VLR molecular valency, b) VLR linker spacing, c) particle size, and d) temperature. We aim to broaden the usefulness of nanoparticle targeting of pathogens with antibodies by 1) identifying design principles that offer a rational method for controlling the molecular affinity through simple modification of valency and 2) utilizing an antibody molecule that is heat-stable up to temperatures of 70¿C or greater. Successful completion of this proposal will open several fruitful paths of discovery which will result in improved sensors and countermeasures for pathogens and biowarfare agents. We anticipate the creation of nano/microparticles with tunable antigen affinity and heat stability. These two impacts will drastically improve the viability of antibody- based detectors, countermeasures, and diagnosis methods. PUBLIC HEALTH RELEVANCE: We propose to create nano/microparticles that are targeted by the newly discovered VLR antibody. We will create recombinant forms of VLRs that we anticipate will offer the following benefits to drug delivery, reagent development, and sensors, including rational adjustment of antibody affinity and temperature stability.

描述（由申请人提供）：我们提出的研究将证明，我们可以通过可变淋巴细胞受体（VLR）抗体的纳米级工程生产出具有可调节亲和力和热稳定性的靶向载体。我们将通过 VLR 功能化纳米/微粒的结合测定来证明这些靶向特性。我们将创建具有一系列化合价、接头长度的多价 VLR 抗体，并且具有适合附着到纳米/微粒上的残基。 VLR 的多价重组形式将识别称为 BclA 的炭疽芽孢杆菌孢子外壳蛋白。我们将通过生物物理技术和单分子成像验证正确的化合价和连接体排列。我们将通过表面等离子共振 (SPR) 和基于原子力显微镜力谱的单分子结合测量来测量 VLR 抗体与 BclA 抗原的结合强度。我们将测量多价抗体的动力学速率和由此产生的亲和力作为价和接头长度的函数。我们还将验证即使在高温下也会发生抗原结合。我们将把多价 VLR 与几种微米/纳米颗粒结合，并使用高通量流式细胞术验证对炭疽芽孢杆菌的粘附。我们将测量颗粒粘附和凝集作为 a) VLR 分子价、b) VLR 连接体间距、c) 颗粒尺寸和 d) 温度的函数。我们的目标是通过以下方式扩大纳米颗粒抗体靶向病原体的用途：1）确定设计原理，提供一种通过简单修饰化合价来控制分子亲和力的合理方法；2）利用热稳定性高达 70°C 或更高的抗体分子。该提案的成功完成将开辟几条富有成效的发现途径，从而改进传感器以及针对病原体和生物战制剂的对策。我们期望创造出具有可调抗原亲和力和热稳定性的纳米/微粒。这两种影响将极大地提高基于抗体的检测器、对策和诊断方法的可行性。 公共健康相关性：我们建议制造新发现的 VLR 抗体靶向的纳米/微粒。我们将创建重组形式的 VLR，预计将为药物输送、试剂开发和传感器带来以下好处，包括合理调整抗体亲和力和温度稳定性。