Secretomes captured in vivo via ultrafiltration probes

通过超滤探针在体内捕获分泌物

• 批准号: 7283959
• 负责人: CHUN-MING HUANG
• 金额: $ 16.82万
• 依托单位: VETERANS MEDICAL RESEARCH FDN/SAN DIEGO
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-09-01 至 2009-08-03
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7283959
• 关键词: Address; Animal Model; Animals; Antibodies; Biological; Biological Models; Blood capillaries; Bypass; Cells; Coupling; Cultured Tumor Cells; Data; Doctor of Philosophy; Enzyme-Linked Immunosorbent Assay; Fiber; Figs - dietary; Glucose; Goals; Immunosuppression; Implant; In Vitro; Ions; Laboratories; Life; Liquid substance; MALDI-TOF Mass Spectrometry; Mass Spectrum Analysis; Membrane; Methods; Modeling; Pattern; Peptides; Property; Proteins; Proteomics; Range; Research; Sampling; System; Techniques; Technology; Therapeutic immunosuppression; Time; Two-Dimensional Gel Electrophoresis; UV induced; Ultrafiltration; Vacuum; animal tissue; base; capillary; cytokine; design; fibrosarcoma; human disease; in vivo; interstitial; killings; neoplastic cell; novel; secretory protein; size; tool; tumor; tumor growth

DESCRIPTION (provided by applicant): It is a challenge to obtain low abundant and/or secretory proteins/peptides (secretomes) from animals in vivo. Our laboratory has developed a novel technique to address this challenge which utilizes capillary ultrafiltration (CUF) probes to capture secretomes in vivo. Capillary ultrafiltration involves a vacuum applied to semipermeable membrane fibers to extract fluid containing secretory molecules from the interstitial space. The CUF probes also provide a simple means to obtain a small-volume sample from living biological systems. We have successfully identified and sequenced the secretomes directly by mass spectrometry, bypassing the traditional proteomics approaches which are primarily based on the 2D electrophoresis gels. In this proposal, we will use the well-established UV-induced fibrosarcoma tumor progressor/regressor model system to exploit the technique of CUF probes to collect in vivo secretomes. Tumor secretomes will be identified using mass spectrometry (MALDI-TOF MS and Q-TOF 2 MS/MS) and enzyme-linked immunosorbent assay (ELISA). In parallel, we will optimize this technique by performing high-throughput mass spectrometry (MALDI-TOF-TOF) and Cytokine Antibody Arrays. We believe that using CUF probes/Mass spectrometry will be a powerful tool which can be applied to other animal models and other human diseases.

描述（由申请人提供）： 从动物体内获取低丰度和/或分泌性蛋白质/肽（分泌组）是一个挑战。我们的实验室开发了一种新技术来应对这一挑战，该技术利用毛细管超滤 (CUF) 探针捕获体内分泌蛋白。毛细管超滤涉及对半透膜纤维施加真空，以从细胞间隙中提取含有分泌分子的液体。 CUF 探针还提供了一种从活生物系统中获取小体积样品的简单方法。我们绕过主要基于二维电泳凝胶的传统蛋白质组学方法，成功地直接通过质谱法对分泌蛋白组进行了鉴定和测序。在本提案中，我们将使用成熟的紫外线诱导纤维肉瘤肿瘤进展/退化模型系统来利用 CUF 探针技术来收集体内分泌组。将使用质谱（MALDI-TOF MS 和 Q-TOF 2 MS/MS）和酶联免疫吸附测定（ELISA）来鉴定肿瘤分泌组。与此同时，我们将通过执行高通量质谱 (MALDI-TOF-TOF) 和细胞因子抗体阵列来优化该技术。我们相信，使用 CUF 探针/质谱将成为一种强大的工具，可应用于其他动物模型和其他人类疾病。