CHEMILUMINESCENT DETECTION OF T CELL PCR PRODUCTS

T 细胞 PCR 产物的化学发光检测

• 批准号: 3489583
• 负责人: KENNETH M KOSAK
• 金额: $ 5万
• 依托单位: LUMITEKK
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-09-01 至 1993-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3489583
• 关键词: DNA; T lymphocyte; alkaline phosphatase; chemical conjugate; chemical reaction; colorimetry; complementary DNA; diagnosis design /evaluation; diagnostic tests; fluoresceins; fluorescence spectrometry; fluorescent dye /probe; gel electrophoresis; hydrogen peroxide; imaging /visualization /scanning; ionophores; light emission; luminescence; method development; nucleic acid chemical synthesis; nucleic acid probes; oxalates; polymerase chain reaction; southern blotting

The polymerase chain reaction (PCR) is a powerful tool used in virtually all aspects of molecular biology. the proposal is to detect PCR products using a primer labeled with a fluorophore that is subsequently activated through a chemiluminescent (CL) reaction. The CL reaction will employ energy transfer, which is capable of activating several fluorophore labels with the same reaction. This has the potential for simultaneously activating and detecting several different colored PCR products in a single step. The additional advantages of nonradioactivity and speed seen with other CL systems should be possible. In this project the primer is labeled with fluorescein. The labeled primer is used to produce the corresponding labeled DNA using PCR. The products are separated by gel electrophoresis and transferred to a membrane. The labeled product is then detected through a CL reaction initiated by the addition of peroxyoxalate and hydrogen peroxide. The light produced gives a direct measurement of the amount of labeled DNA present. If successful, different colored PCR products could be detected simultaneously in the same preparation. Colored CL testing could detect multiple DNA probes for rapid screening or diagnostic testing.

聚合酶链反应（PCR）是一种实用的强大工具 分子生物学的所有方面。 该建议是检测PCR产品 使用用荧光团标记的底漆，随后被激活 通过化学发光（CL）反应。 CL反应将采用 能量转移，能够激活多个荧光团标签 具有相同的反应。 这有可能同时 激活和检测单个彩色PCR产品 步。 非放射性和速度的其他优势 其他CL系统应该是可能的。 在此项目中，底漆用荧光素标记。 标记的底漆 用于使用PCR产生相应的标记DNA。 产品 通过凝胶电泳分离并转移到膜上。 这 然后通过由由CL反应检测到标记的产物 添加过氧化氧化物和过氧化氢。 产生的光给 直接测量存在标记的DNA的量。 如果成功，可以检测到不同颜色的PCR产品 同时在相同的准备工作中。 彩色CL测试可以检测到 多个DNA探针进行快速筛查或诊断测试。