RBC typing antibodies labeled with rare earth phosphor upconverting nanocrystals

描述（由申请人提供）：Sunstone Biosciences和宾夕法尼亚大学提出的STTR申请将产生的产品是一种新型标记抗体为基础的试剂盒，可用于在输血前快速识别各种红细胞表型。目前，输血前血型仅涉及ABO和Rh组的主要抗原，并且主要依赖于人工血凝反应的使用。该程序的时间和劳动强度限制了输血前可以检测的RBC亚群的数量，有可能使一生中多次输血的患者发生致敏。我们计划开发一个大型的红细胞抗原特异性单克隆抗体，用稀土纳米荧光粉标记，能够快速确定红细胞表型，使用广泛的常见和罕见抗原，自动化，多路分析格式。稀土晶体的可编程和窄光谱是产生分子检测系统的理想选择，它比传统的光学探针具有更大的多路复用能力。在计划的第二阶段结束时，我们预计将设计并构建一个由大约20个或更多抗体组成的小组，每个抗体都具有独特的光学发射信号，可用于创建革命性的血液表型“特征”。纳米晶体的上转换特性是靶向标记和定量的唯一标识。其他分子检测方法，包括量子点和传统荧光团，在整个研究界被广泛使用，但具有许多局限性，如光漂白，分辨率差，以及允许同时运行的样品数量有限。稀土荧光粉纳米晶体解决了这些问题，作为惰性的、稳定的光发射器，可以实现高分辨率和广泛的多路成像。公共卫生相关性：Sunstones Biosciences与其在普林斯顿大学的合作伙伴已经产生了初步证据，证明可以合成小至~ 10nm的纳米晶体，这些纳米晶体保留其独特的光学特性，并且可以被涂覆以实现功能化（例如与抗体结合）。在这个第一阶段的提案中，我们将扩展这些研究，以产生和测试至少三种荧光纳米晶体标记的红细胞抗原特异性抗体。该提案将通过开发一种全面匹配供体和受体血型的新产品，减少过敏反应和其他输血副作用的发生率。与传统的血液分型方法不同，该方法将允许快速检测甚至罕见的血液标记物。

DESCRIPTION (provided by applicant): The product that will result from the proposed STTR application from Sunstone Biosciences and the University of Pennsylvania is a novel labeled antibody-based reagent kit that can be used to rapidly identify diverse red blood cell phenotypes prior to blood transfusion. At present, pre-transfusion blood typing involves only the major antigens of the ABO and Rh groups, and relies on the use of a largely- manual hemagglutination reaction. The time- and labor-intensiveness of this procedure limits the number of RBC sub-groups that can be tested prior to a transfusion, risking the development of sensitization of patients who receive multiple transfusions throughout their life. We plan to develop a large panel of RBC antigen-specific MAbs, labeled with rare-earth nanophosphors, able to rapidly determine RBC phenotype using a wide range of common and rare antigens in an automatable, multiplexed assay format. The programmable, and narrow optical spectra of rare-earth crystals are ideal for generating molecular detection systems that are capable of vastly greater multiplexing than conventional optical probes. At the end of Phase 2 of the proposed program, we anticipate having designed and constructed a panel of approximately 20 or more antibodies, each with a unique optical emission signal, that can be used to create a revolutionary blood phenotyping "signature". The upconverting characteristics of the nanocrystals are unique identifiers of target-specific labeling and quantification. Other methods of molecular detection, including Quantum Dots and conventional fluorophores, are utilized widely throughout the research community but possess many limitations such as photobleaching, poor resolution, and the limited number of samples permitted to run simultaneously. Rare-earth phosphor nanocyrstals address these problems, as inert, stable light emitters that allow for high resolution and vastly multiplexed imaging. PUBLIC HEALTH RELEVANCE: Sunstones Biosciences has, with its consortium partners at Princeton University, generated preliminary evidence that nanocrystals as small as ~10 nm can be synthesized, that these nanocrystals retain their unique optical properties, and that they can be coated for functionalization (such as for conjugation to antibodies). In this Phase 1 proposal, we will extend these studies to generate and test at least three phosphor nanocrystal-labeled antibodies specific for RBC antigens. This proposal will reduce the incidence of allergic reactions and other side-effects of blood transfusion by developing a new product for comprehensively matching donor and recipient blood types. Unlike conventional methods for blood typing, the proposed method will allow rapid detection of even rare blood markers.