The Transfusion Chip: A Simple, Low Cost Microarray for DNA Based Blood Typing

描述（申请人提供）：低成本的核酸分析，检测HIV和HBV，彻底改变了血液供应的筛选。尽管有了这种新的遗传复杂性，传统的血型分型[ABO， Rh]在很大程度上仍然是通过80年前的凝集试验进行的。最近，产生标准血型的基因变异已经被确定，并显示出相对简单。同时，有研究表明，为了提高输血质量，历史标记集可能会扩大到包括更完整的7个特征组：[ABO， Rh, Duffy, Kidd, Kell, Dombrock & MNS]，这些特征的潜在遗传学现在已经知道。这两种平行的趋势表明，基因检测已经成为血型筛查的未来：但前提是基因检测的成本和技术简单程度与凝集或目前的核酸病原体检测大致相同。我们观察到，只要稍加修改，我们为基于原始血液的微阵列hla分型开发的生化、硬件和软件技术套件就可以转化为低成本的血型分型技术，我们称之为“输血芯片”。在这个为期6个月的第一阶段计划中，我们提出了2个具体目标作为里程碑，以证明随后的第二阶段的合理性。SA1。设计并初步验证13-15个PCR反应并行进行。7个血液标记基因中的所有信息位点将被平行扩增，以产生一个染料标记的扩增子集，准备用于微阵列杂交。SA2。微阵列探针的设计，制造和初步验证。一个输血芯片原型将被设计和制造，以询问信息性的序列变异，这些序列变异定义了7个位点之间的血型变异。验证将使用来自SA1的扩增dna。与后续第二阶段的关系。第一阶段的完成将产生基于pcr的样品标记反应原型（适用于原始血液）和用于DNA血型的低成本输血芯片微阵列原型。第二阶段将专注于改进PCR和微阵列设计，扩大生产规模，初步PCR试剂盒制造和将这些材料交付给beta测试者，包括3-4个顶级血液实验室：将输血芯片的性能与血清学分型以及与之竞争的微阵列和PCR测试进行比较。公共卫生叙事。病原体污染的基因检测已经彻底改变了血液供应的安全性，但由于成本和复杂性，类似的基于DNA的潜在血型检测已经落后。我们在这里提出的一种基于微阵列的血型分型方法，将允许血型分析现代化，通过利用二十年来血型遗传学的价值：允许血型抗原变异的分析，以一种比血清学更完整的方式获得，在成本和技术简单程度上与血液病原体筛选一样实用。由此产生的技术不仅将降低美国血液供应检测的成本，而且我们认为，将使发展中国家的新一代增强的、基于dna的血型分型成为可能。

DESCRIPTION (provided by applicant): Low-cost nucleic acid analysis, to detect HIV & HBV, has revolutionized screening of the blood supply. In spite of that new genetic sophistication, traditional blood group typing [ABO, Rh] is still performed, for the most part, via variations upon 80-year-old agglutination testing. Recently, the genetic variation which gives rise to the standard blood groups has been defined, and shown to be relatively simple. In parallel, it has been shown that, to enhance transfusion quality, the historical marker set might be expanded to include a more complete panel of 7 traits: [ABO, Rh, Duffy, Kidd, Kell, Dombrock & MNS] for which the underlying genetics are now known. These two parallel trends suggest that genetic testing has emerged as the future of blood group screening: but only if the genetic test can be delivered at roughly the same cost and level of technical simplicity as agglutination, or the current panel of nucleic acid based pathogen tests. We have observed that, with minor modification, the suite of biochemical, hardware & software technologies that we have developed for microarray based HLA-typing on raw blood could, instead, be transformed into low-cost technologies for blood-group typing, which we refer to as "The Transfusion-Chip". In this 6-month Phase I plan, we propose 2 Specific Aims as Milestones to justify a subsequent Phase II. SA1. Design & preliminary validation of 13-15 PCR reactions to be performed in parallel. All informative sites among the set of 7 blood marker genes will be amplified in parallel, to generate a dye-labeled amplicon set that is ready to be used for microarray hybridization. SA2. Microarray probe design, fabrication &preliminary validation. A Transfusion-Chip prototype will be designed and fabricated, to interrogate the informative sequence variations which define blood group variation among the set of 7 loci. The validation will employ amplified DNAs from SA1. Relation to a Follow-on Phase II. Completion of Phase I will yield a prototype PCR-based sample labeling reaction (that works on raw blood) and a prototype low-cost Transfusion-Chip microarray for DNA based blood-typing. Phase II will focus on refinement of the PCR and microarray designs, manufacturing scale-up, preliminary PCR kit fabrication and delivery of those materials to beta testers, comprising 3-4 top blood labs: to compare Transfusion-Chip performance to serological typing and to the competing microarray and PCR tests. Public Health Narrative. Genetic testing for pathogen contamination has revolutionized the safety of the blood supply, yet analogous DNA based testing of the underlying blood groups has lagged-behind, due to cost and complexity. A microarray-based approach to blood group typing that we propose here, will allow blood type analysis to be modernized, by exploiting two decades worth of blood group genetics: allowing analysis of blood group antigen variation, in a way that is more complete than can be obtained by serology, at a cost and level of technical simplicity that is as practical as that of blood pathogen screening. The resulting technology will not only reduce the cost of blood supply testing in the US, but we propose, will enable a new generation of enhanced, DNA-based blood group typing in the developing world. PUBLIC HEALTH RELEVANCE: A microarray-based approach to blood group typing that we propose here, will allow blood type analysis to be modernized, by exploiting two decades worth of blood group genetics: allowing analysis of blood group antigen variation, in a way that is more complete than can be obtained by serology, at a cost and level of technical simplicity that is as practical as that of blood pathogen screening. The resulting technology will not only reduce the cost of blood supply testing in the US, but we propose, will enable a new generation of enhanced, DNA-based blood group typing in the developing world.