A Novel Multiplexed Assay for Rapid Antibody Screening

一种用于快速抗体筛选的新型多重检测方法

• 批准号: 8314092
• 负责人: Scott Allen Monsma
• 金额: $ 38.36万
• 依托单位: PRIMORIGEN BIOSCIENCES, INC.
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-09-01 至 2014-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8314092
• 关键词: Address; Algorithms; Amino Acid Sequence; Amylases; Antibodies; Antigens; Beta Cell; Biological Assay; Biological Markers; Cell Culture Techniques; Cell Differentiation process; Cell Line; Cell Maturation; Cell Therapy; Cells; Cessation of life; Characteristics; Chinese Hamster Ovary Cell; Computer software; Custom; Data; Data Analyses; Data Reporting; Detection; Development; Endocrine; Endoderm; Enzyme-Linked Immunosorbent Assay; Enzymes; Factor Analysis; Funding; Generations; Genes; Glucagon; Glucokinase; Goals; Growth; Guidelines; Hormones; Hour; Human; Human Genome; Image; Immunoassay; In Vitro; Individual; Insulin; Insulin-Dependent Diabetes Mellitus; Islet Cell; Legal patent; Letters; Linear Regressions; Marketing; Metabolism; Methods; Molecular Profiling; Mus; Nuclear; Oryctolagus cuniculus; Pancreas; Pancreatic Polypeptide; Pattern; Peptide Sequence Determination; Performance; Phase; Population; Proteins; Proteome; Rattus; Reagent; Recombinant Proteins; Reference Standards; Regenerative Medicine; Regulation; Reproduction; Research; Research Personnel; Run-On Assays; Sampling; Screening procedure; Small Business Innovation Research Grant; Solutions; Somatostatin; Specificity; Spottings; Staging; Stem cells; Structure of beta Cell of islet; System; Testing; Time; Touch sensation; Tumor Cell Line; Universities; Validation; Variant; Wisconsin; assay development; base; cell type; cellular development; computerized data processing; cost; data acquisition; design; embryonic stem cell; extracellular; high throughput screening; induced pluripotent stem cell; internal control; islet; medical schools; miniaturize; novel; phase 1 study; phase 2 study; polyclonal antibody; progenitor; programs; prototype; public health relevance; research study; software development; synthetic peptide; therapy development; tool; transcription factor; user friendly software

DESCRIPTION (provided by applicant): Primorigen Biosciences will use Phase II SBIR funds to accelerate and expand the development of new regenerative medicine cell therapies by developing a novel high throughput, low cost protein quantification system for characterizing pancreatic 2 cell and islet cell differentiation. Primorigen successfully confirmed the technical feasibility of this proposal by achieving both major goals of the Phase I project, first demonstrating (and now routinely using) its proprietary Spots on Dots" frameless microarray platform for high throughput antibody screening and characterization, and then demonstrating feasibility of developing a multiplexed protein quantitation assay for pancreatic beta cell differentiation by producing a prototype multiplexed assay with matched antibody pairs for quantifying two important markers of 2 cell differentiation (Nkx2.2 and MafB) and with 9 additional assays (Amylase 2B, FoxA2, GCK, HNF6, Isl1, NeuroD1, Neurog3, Pax4 and Pax6) in advanced stages of testing and development. In Phase II, Primorigen will complete the existing panel of assays for up to 11 markers of 2 cell differentiation (Phase I) and develop a new panel for identifying and characterizing mature islet cell populations, including 1, 2, 4, 5, and Pancreatic Polypeptide (PP) cell types. These combined tools will provide a more complete solution for tracking downstream development and maturation of pancreatic beta cells, and will be validated by obtaining independent corroboration of the performance and specificity of the assays. In addition, Primorigen will collaborate with a software developer to produce a software/flatbed scanner package that enables standardized analysis and reporting of data from assays run on the Spots on Dots" platform in a rapid, 'one touch' format. PUBLIC HEALTH RELEVANCE: Proteins facilitate many of the cell's most basic functions of reproduction, metabolism, growth, and programmed death. In the past few years, thousands of new protein sequences and post- translational variants have been identified by the human genome and proteome projects. This protein jackpot has hastened the drive to understand protein-based regulation but it also challenges researchers to find better biophysical methods to quantitatively detect protein markers. As a result, there is demand for inexpensive, miniaturized, multiplexed high throughput assays that can generate thousands of protein detection data points per experiment. Primorigen's SBIR proposal will address this need by developing well-characterized antibody and protein assay content. The assays will be valuable for detecting and quantifying markers of pancreatic beta- and islet-cell differentiation and maturation. The assays will be formatted in a high throughput, low-cost multiplexed platform that is supported by one touch user friendly software for data analysis and reporting.

描述（由申请人提供）：Primorigen Biosciences将使用第二阶段SBIR资金，通过开发一种用于表征胰腺2细胞和胰岛细胞分化的新型高通量、低成本蛋白质定量系统，加速和扩大新的再生医学细胞疗法的开发。Primorigen通过实现第一阶段项目的两个主要目标，成功地证实了该提案的技术可行性，（现在常规使用）其专有的点上点”无框架微阵列平台，用于高通量抗体筛选和表征，然后通过产生具有以下的原型多重测定来证明开发用于胰腺β细胞分化的多重蛋白质定量测定的可行性用于定量2细胞分化的两种重要标志物（Nkx2.2和MafB）的匹配抗体对，以及在测试和开发的后期阶段的9种额外测定（淀粉酶2B、FoxA 2、GCK、HNF 6、Isl 1、NeuroD 1、Neurog 3、Pax 4和Pax 6）。在第二阶段，Primorigen将完成现有的2种细胞分化的11种标志物的检测（第一阶段），并开发一种新的检测组，用于鉴定和表征成熟胰岛细胞群，包括1，2，4，5和胰腺多肽（PP）细胞类型。这些组合工具将为跟踪胰腺β细胞的下游发育和成熟提供更完整的解决方案，并将通过获得检测性能和特异性的独立确证进行验证。此外，Primorigen将与一家软件开发商合作，开发一个软件/平板扫描仪包，以快速、“一触式”的格式对在Spots on Dots平台上运行的检测数据进行标准化分析和报告。 公共卫生相关性：蛋白质促进细胞的许多最基本的功能，如生殖、代谢、生长和程序性死亡。在过去的几年中，人类基因组和蛋白质组计划已经鉴定了数千种新的蛋白质序列和翻译后变体。这种蛋白质头奖加速了理解基于蛋白质的调控的动力，但它也挑战研究人员找到更好的生物物理方法来定量检测蛋白质标记。因此，需要廉价的、小型化的、多路复用的高通量测定，其可以在每个实验中产生数千个蛋白质检测数据点。Primorigen的SBIR提案将通过开发良好表征的抗体和蛋白质检测内容来满足这一需求。该测定法对于检测和定量胰腺β细胞和胰岛细胞分化和成熟的标志物将是有价值的。将在一个高通量、低成本的多路复用平台中对检测进行格式化，该平台由一触式用户友好软件支持，用于数据分析和报告。