Ultra high sensitivity mix and measure immunoassays in blood

描述（由申请人提供）：MagneSensors的I期SBIR在新的超灵敏三明治免疫分析中使用磁传感器和磁性纳米颗粒标签。重点是使用非常理想的混合和测量格式测量血液中非常低水平的抗原。这种组合极难实现，目前在竞争的生物测定平台上无法获得。磁检测具有独特的优势，如结合的定量测量，而不需要通常分离未结合的标签，以及血液和血清中固有的缺乏磁背景。该磁检测平台利用高温超导量子干涉器件（squid）检测磁性纳米颗粒标记的抗体与抗原的特异性结合。重要的是，三明治免疫测定的超高灵敏度将利用我们的新一代磁性检测仪器，该仪器将很快在NIH的一个单独项目下上线。这种新的台式仪器使用一个小型冷冻冰箱，提供比我们现有仪器更好的50-200倍的信噪比，以及更高的吞吐量。高灵敏度提供了对小体积有价值试剂或稀缺样品的早期检测和测量的优势。混合和测量格式显着减少了分析准备时间，简化了自动化（无洗涤步骤），从而实现了快速，高通量的分析。在血液和血清中起作用的能力为更广泛的重要免疫测定带来了这些好处。第一阶段的目标是在白细胞介素-6 （IL-6）模型系统上演示超高灵敏度磁夹心免疫分析，目标如下：1)全血，2)<0.4 pg/ml, 3)不分离未结合的磁性纳米颗粒标记，4)包括制备和测量在内的总分析时间<60分钟。第一阶段的重点是使用夹心磁免疫分析法检测血液中的抗原。0.4 pg/ml的检测目标是血液中内源性IL-6水平的下限。第一阶段将使用我们开发的一种新技术来实现，该技术使用二氧化硅微珠和密度梯度介质在磁检测之前从血细胞中去除捕获的抗原。需要注意的是，第一阶段的工作并没有在血液中开发出真正的混合和测量免疫测定，因为在第一阶段，血细胞从捕获的抗原中分离出来。这个更雄心勃勃的目标，目前在其他检测平台的超高灵敏度下无法实现，将留到第二阶段。

DESCRIPTION (provided by applicant): MagneSensors' Phase I SBIR uses magnetic sensors and magnetic nanoparticle labels in new ultra-sensitive sandwich immunoassays. The focus is on measuring very low levels of antigens in blood using the highly desirable mix and measure format. This combination is extremely difficult to achieve and is currently not available in competing bioassay platforms. Magnetic detection offers unique advantages, such as the quantitative measurement of binding without requiring the usual separation of unbound labels, as well as the inherent lack of magnetic background in blood and serum. This magnetic detection platform uses high temperature superconducting quantum interference devices (SQUIDs) to detect the specific binding to antigens of antibodies labeled with magnetic nanoparticles. Importantly, the ultra-high levels of sensitivity for sandwich immunoassays will take advantage of our new, next-generation magnetic detection instrument that will be brought on line shortly under a separate NIH program. This new bench top instrument uses a small cryorefrigerator and offers 50-200X better signal-to-noise than our existing instrument along with much higher throughput. High sensitivity offers advantages of early detection and measurements on small volumes of valuable reagents or scarce samples. The mix and measure format significantly reduces assay preparation time and simplifies automation (no wash steps), thereby enabling rapid, high throughput assays. The ability to work in blood and serum brings these benefits to a much broader range of important immunoassays. The Phase I aim is to demonstrate ultra high sensitivity magnetic sandwich immunoassays on an interleukin-6 (IL-6) model system with the following goals: 1) whole blood, 2) <0.4 pg/ml, 3) no separation of unbound magnetic nanoparticle labels, 4) <60 minutes total assay time including preparation and measurement. Phase I focuses on detection of an antigen in blood using a sandwich magnetic immunoassay format. The detection goal of 0.4 pg/ml is at the lower end of endogenous levels of IL-6 in blood. It will be achieved in Phase I using a novel technique we have developed that uses silica microbeads and a density gradient media to remove the captured antigen from blood cells prior to magnetic detection. Note that the Phase I effort does not develop a true mix and measure immunoassay in blood, as blood cells are separated out from captured antigen in Phase I. This more ambitious goal, currently unachievable at ultra-high sensitivity with other detection platforms, is left for Phase II.