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1. 发明授权

US07202043B2 Detecting cell membrane protein endocytosis 有权
标题翻译：检测细胞膜蛋白内吞
公开(公告)号：US07202043B2
公开(公告)日：2007-04-10
申请号：US11244256
申请日：2005-10-04
申请人： Yun-Jung Choi , Kun Peng , Linda Kauffman , Rajendra Singh
发明人： Yun-Jung Choi , Kun Peng , Linda Kauffman , Rajendra Singh
IPC分类号： G01N33/53
CPC分类号： C12Q1/37 , G01N33/5005 , G01N33/567 , G01N2333/924 , G01N2333/976 , Y10S435/962 , Y10S435/963
摘要： Methods and reagents are provided for determining endocytosis, using a cell expressing an externally polypeptide labeled cell membrane receptor, and as a reagent an antibody to said label conjugated to a fragment of an enzyme fragment complementation pair. Compounds are tested for their effect on endocytosis by complexing the reagent with said cells, adding the test compound at any time in relation to the complexing, allowing any endocytosis to occur, proteolytically degrading external enzyme fragment, adding protease inhibitor and the complementary member of the enzyme fragment complementation pair and substrate. The product provides a detectable signal related to the amount of endocytosis that occurred. The method is readily automated as all steps can occur in a single vessel without separations and washings.
摘要翻译：提供了用于确定内吞作用的方法和试剂，使用表达外部多肽标记的细胞膜受体的细胞，以及作为与所述酶标记片段互补对的片段缀合的所述标记的抗体作为试剂。通过使试剂与所述细胞络合来测试化合物对内吞作用的影响，在任何时间加入测试化合物与复合相关，允许发生任何内吞作用，蛋白水解降解外部酶片段，加入蛋白酶抑制剂和补体成员酶片段互补对和底物。该产品提供与发生的胞吞作用量相关的可检测信号。该方法易于自动化，因为所有步骤都可以发生在没有分离和洗涤的单个容器中。

2. 发明申请

US20060073523A1 Detecting cell membrane protein endocytosis 有权
标题翻译：检测细胞膜蛋白内吞
公开(公告)号：US20060073523A1
公开(公告)日：2006-04-06
申请号：US11244256
申请日：2005-10-04
申请人： Yun-Jung Choi , Kun Peng , Linda Kauffman , Rajendra Singh
发明人： Yun-Jung Choi , Kun Peng , Linda Kauffman , Rajendra Singh
IPC分类号： G01N33/567 , G01N33/53
CPC分类号： C12Q1/37 , G01N33/5005 , G01N33/567 , G01N2333/924 , G01N2333/976 , Y10S435/962 , Y10S435/963
摘要： Methods and reagents are provided for determining endocytosis, using a cell expressing an externally polypeptide labeled cell membrane receptor, and as a reagent an antibody to said label conjugated to a fragment of an enzyme fragment complementation pair. Compounds are tested for their effect on endocytosis by complexing the reagent with said cells, adding the test compound at any time in relation to the complexing, allowing any endocytosis to occur, proteolytically degrading external enzyme fragment, adding protease inhibitor and the complementary member of the enzyme fragment complementation pair and substrate. The product provides a detectable signal related to the amount of endocytosis that occurred. The method is readily automated as all steps can occur in a single vessel without separations and washings.
摘要翻译：提供了用于确定内吞作用的方法和试剂，使用表达外部多肽标记的细胞膜受体的细胞，以及作为与所述酶标记片段互补对的片段缀合的所述标记的抗体作为试剂。通过使试剂与所述细胞络合来测试化合物对内吞作用的影响，在任何时间加入测试化合物与复合相关，允许发生任何内吞作用，蛋白水解降解外部酶片段，加入蛋白酶抑制剂和补体成员酶片段互补对和底物。该产品提供与发生的胞吞作用量相关的可检测信号。该方法易于自动化，因为所有步骤都可以发生在没有分离和洗涤的单个容器中。

3. 发明申请

US20060003388A1 Sensitive intracellular calcium assay 有权
标题翻译：敏感的细胞内钙测定
公开(公告)号：US20060003388A1
公开(公告)日：2006-01-05
申请号：US11135779
申请日：2005-05-23
申请人： Linda Kauffman , Rajendra Singh , Edwin Ullman
发明人： Linda Kauffman , Rajendra Singh , Edwin Ullman
IPC分类号： G01N33/53
CPC分类号： G01N33/582
摘要： A sensitive intracellular calcium assay is disclosed comprising conveniently a reagent comprised of a dye precursor capable of entering cells and being hydrolyzed to a dye, whereby the dye complexes with calcium in said cells and provides a luminescent signal, an antibody specific for the dye and conjugated with a quencher, and a cellular anion exchange enzyme inhibitor. In performing the assay, the reagent is combined with cells expressing a receptor responsive to a ligand resulting in a change in cytosolic calcium. After incubation for the dye precursor to permeate the cells, the calcium may be determined by exciting the dye precursor and determining the peak fluorescence over a time course. The method can be used for measuring the effect of an agent on cytosolic calcium by binding to a cell surface membrane receptor.
摘要翻译：公开了敏感的细胞内钙测定法，其包括方便的是由能够进入细胞并被水解成染料的染料前体组成的试剂，由此所述染料与所述细胞中的钙络合并提供发光信号，对于染料和缀合的抗体与猝灭剂和细胞阴离子交换酶抑制剂。在进行测定时，试剂与表达对配体有影响的受体的细胞结合，导致细胞溶质钙的变化。在孵育染料前体渗透细胞后，可以通过激发染料前体并在时间过程中确定峰荧光来确定钙。该方法可用于通过结合细胞表面膜受体来测量药剂对细胞溶质钙的影响。

4. 发明授权

US07371534B2 Sensitive intracellular calcium assay 有权
标题翻译：敏感的细胞内钙测定
公开(公告)号：US07371534B2
公开(公告)日：2008-05-13
申请号：US11135779
申请日：2005-05-23
申请人： Linda Kauffman , Rajendra Singh , Edwin F. Ullman
发明人： Linda Kauffman , Rajendra Singh , Edwin F. Ullman
IPC分类号： G01N33/53 , A61K51/00
CPC分类号： G01N33/582
摘要： A sensitive intracellular calcium assay is disclosed comprising conveniently a reagent comprised of a dye precursor capable of entering cells and being hydrolyzed to a dye, whereby the dye complexes with calcium in the cells and provides a luminescent signal, an antibody specific for the dye and conjugated with a quencher, and a cellular anion exchange enzyme inhibitor. In performing the assay, the reagent is combined with cells expressing a receptor responsive to a ligand resulting in a change in cytosolic calcium. After incubation for the dye precursor to permeate the cells, the calcium may be determined by exciting the dye precursor and determining the peak fluorescence over a time course. The method can be used for measuring the effect of an agent on cytosolic calcium by binding to a cell surface membrane receptor.
摘要翻译：公开了敏感的细胞内钙测定法，其包括方便的是由能够进入细胞并被水解成染料的染料前体组成的试剂，由此染料与细胞中的钙络合并提供发光信号，对于染料和缀合的抗体与猝灭剂和细胞阴离子交换酶抑制剂。在进行测定时，试剂与表达对配体有影响的受体的细胞结合，导致细胞溶质钙的变化。在孵育染料前体渗透细胞后，可以通过激发染料前体并在时间过程中确定峰荧光来确定钙。该方法可用于通过结合细胞表面膜受体来测量药剂对细胞溶质钙的影响。

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