Chromatographic Automation of Immunoassays

描述（由申请人提供）：临床和药物化学家对血液或血清中许多药物和激素的游离或非结合部分非常感兴趣，因为这些部分被认为代表了许多此类药物的活性形式。因此，这些药物和激素的游离部分应该作为患者诊断和治疗的理想工具进行测量。然而，目前还没有通用和快速的方法来测量临床样品中药物和激素的游离部分。本提案的总体目标是：(1)更好地了解药物和激素与血液或血清中的结合剂的相互作用；(2)利用这些信息开发快速、可靠的游离药物和激素检测。该项目的基本假设是，基于超快速亲和萃取和层析免疫分析技术，可以创建一类新的改进的游离药物和激素检测方法，如最近对华法林、甲状腺素、苯妥英和卡马西平等化合物的检测。未来的研究将建立在这些以前的努力，考虑新的分析和方法，可以在这样的工作中使用。特别强调将给予高通量的方法和技术，可以适用于广泛的分析物的检测。结合研究领域有待探索的进展将包括开发快速亲和方法，用于检查药物或激素与血清药物相互作用的程度和速率，以及使用亲和微柱作为个性化医疗工具，研究药物与血清蛋白变体相互作用的变化。用于游离药物和激素测量的改进工具将包括用于快速药物或激素提取的超高容量亲和支架，用于柱上检测的色谱免疫测定，用于游离药物/激素测定的多维系统，使用血清蛋白柱进行游离药物/激素测定，改进了分析平台，提高了近红外荧光标记层析免疫测定的灵活性和样品通量。这项工作的直接影响将是更好地了解药物和激素如何与血清结合剂相互作用并在循环系统中运输。此外，用于自由药物/激素测量的新技术将大大提高临床医生和药物化学家研究和检查体内药物和激素的生物效应的能力。这种能力将导致临床试验和个性化医疗的更好方案，以治疗和诊断患有各种疾病的患者。在这个项目中开发的各种基于亲和的方法，用于检查溶质配体结合，从生物样品中快速提取分析物，以及测量这些分析物的痕量水平，在生物化学研究的许多领域，包括蛋白质组学，代谢组学和候选药物的表征或筛选，也应该是有价值的。

DESCRIPTION (provided by applicant): The free, or non-bound, fraction of many drugs and hormones in blood or serum is of great interest to clinical and pharmaceutical chemists because this fraction is believed to represent the active form of many such agents. As a result, it is the free fraction of these drugs and hormones that should ideally be measured as a tool for patient diagnosis and treatment. However, there is currently no general and fast approach for measuring the free fractions of drugs and hormones in clinical samples. The overall goals of this proposal are 1) to obtain a better understanding of drug and hormone interactions with their binding agents in blood or serum and 2) to use this information to develop fast, reliable free drug and hormone assays. The underlying hypothesis of this project is that a new class of improved free drug and hormone assays can be created based on the techniques of ultrafast affinity extraction and chromatographic immunoassays, as recently demonstrated for compounds such as warfarin, thyroxine, phenytoin and carbamazepine. Future studies will build on these previous efforts by considering new analytes and approaches that can be employed in such work. Particular emphasis will be given to high-throughput methods and techniques that can be adapted for the detection of a broad range of analytes. Advances to be explored in the field of binding studies will include the development of rapid affinity methods for examining both the extent and rate of drug or hormone interactions with serum agents, and the use of affinity microcolumns as tools in personalized medicine to study changes in drug interactions with variants of serum proteins. Improved tools to be created for free drug and hormone measurements will include ultra-high capacity affinity supports for use in fast drug or hormone extractions, chromatographic immunoassays with on-column detection, multi-dimensional systems for free drug/hormone assays, the use of serum protein columns to carry out free drug/hormone assays, and improved analytical platforms for improving the flexibility and sample throughput when using chromatographic immunoassays with near infrared fluorescent labels. An immediate impact of this work will be a better understanding of how drugs and hormones interact with serum binding agents and are transported in the circulatory system. In addition, the new techniques that will be created for free drug/hormone measurements should greatly improve the ability of clinicians and pharmaceutical chemists to study and examine the biological effects of drugs and hormones in the body. This capability should lead to better protocols in clinical testing and in personalized medicine for the treatment and diagnosis of patients suffering from a variety of disorders. The various affinity-based approaches developed in this project for examining solute-ligand binding, rapidly extracting analytes from biological samples, and measuring trace levels of such analytes should also be valuable in many areas of biochemical research, including proteomics, metabolomics and the characterization or screening of drug candidates.