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31. 发明授权

US07108996B2 Method for increasing luminescence assay sensitivity 有权
标题翻译：增加发光测定灵敏度的方法
公开(公告)号：US07108996B2
公开(公告)日：2006-09-19
申请号：US10991759
申请日：2004-11-18
申请人： Erika Hawkins , John M. Centanni , Jacqueline Sankbeil , Keith V. Wood
发明人： Erika Hawkins , John M. Centanni , Jacqueline Sankbeil , Keith V. Wood
IPC分类号： C12Q1/66
CPC分类号： C12Q1/66 , G01N33/582 , G01N2333/90241 , Y10S435/968 , Y10S435/975
摘要： A method for increasing the sensitivity of a bio-luminescent assay comprising carrying out the assay in the presence of an organic compound that reduces luminescence that is not dependent on the presence of an analyte by at least about 10 fold, and that reduces, maintains, or increases the luminescence that is dependent on the presence of an analyte.
摘要翻译：一种用于增加生物发光测定的灵敏度的方法，包括在有机化合物存在下进行测定，所述有机化合物将不依赖于分析物的存在的发光减少至少约10倍，并且降低，维持，或增加取决于分析物存在的发光。

32. 发明授权

US5814471A Luciferase inhibitor compositions and methods of using same 失效
标题翻译：荧光素酶抑制剂组合物及其使用方法
公开(公告)号：US5814471A
公开(公告)日：1998-09-29
申请号：US831781
申请日：1997-04-09
申请人： Keith V. Wood
发明人： Keith V. Wood
IPC分类号： G01N21/76 , C12N9/02 , C12Q1/66 , G01N33/53 , H01L21/306
CPC分类号： C12N9/0069 , C12Q1/66 , C12Y113/12007 , Y10S435/81 , Y10S435/968 , Y10S435/975 , Y10S436/808
摘要： Methods and compositions are provided for improved kinetics of light production from luciferase activity in beetle luciferase-luciferin reactions utilizing a luciferase inhibitor. Thus, the invention provides methods and compositions for assaying samples for the presence of a beetle luciferase in conjunction with a luciferase inhibitor.
摘要翻译：提供了方法和组合物，用于利用荧光素酶抑制剂改善甲虫荧光素酶 - 荧光素反应中荧光素酶活性的光产生动力学。因此，本发明提供了用于测定样品与荧光素酶抑制剂结合的甲虫荧光素酶的存在的方法和组合物。

33. 发明授权

US5583024A Recombinant expression of Coleoptera luciferase 失效
标题翻译：鞘翅目萤光素酶的重组表达
公开(公告)号：US5583024A
公开(公告)日：1996-12-10
申请号：US60091
申请日：1993-05-10
申请人： Marlene D. McElroy, deceased , W. D. McElroy, executor , Donald R. Helinski , Keith V. Wood , Jeffrey R. De Wet , David W. Ow , Stephen H. Howell
发明人： Marlene D. McElroy, deceased , W. D. McElroy, executor , Donald R. Helinski , Keith V. Wood , Jeffrey R. De Wet , David W. Ow , Stephen H. Howell
IPC分类号： C12N1/21 , C12N9/02 , C12N9/38 , C12N15/52 , C12Q1/68 , G01N33/535 , G01N33/573 , C12N15/53 , C12N15/63
CPC分类号： C12N9/0069 , C12N15/52 , C12Q1/6897 , C12Y113/12007 , C12Y302/01023 , G01N33/535 , G01N33/573 , C07K2319/00 , C07K2319/61
摘要： A method is disclosed for producing a protein which expresses bioluminescence activity which involves combining two polydeoxyribonucleotides, one containing a continuous sequence of codons encoding a polypeptide which comprises a single covalently bonded molecular structure and which catalyzes the oxidation of insect luciferin to yield light and the other which causes DNA transcription, and obtaining the polypeptide by transcription and subsequent translation. The insect luciferin is derived from bioluminescent insect, preferably Diptera and Coleoptera (fireflies and beetles). Hybrid proteins are similarly formed by inclusion of an additional polydeoxyribonucleotide encoding for a second polypeptide such that their respective polypeptide-encoding reading frames form a continuous reading frame. Also disclosed is a method for quantitatively assaying a fluid for the presence of an unknown quantity of antigen using the hybrid proteins bonded to antibody directed against the antigen, determining luminescence after reaction of the antibody and antigen, and determining the amount of the antigens by comparison of the determined luminescence with the luminescence previously measured under equivalent conditions for equivalent hybrid proteins reacted under equivalent conditions with known amounts of the antigen.
摘要翻译：公开了用于产生表达生物发光活性的蛋白质的方法，其涉及组合两种多脱氧核糖核苷酸，一种含有编码多肽的密码子的连续序列，其包含单个共价键合的分子结构并且催化昆虫萤光素的氧化以产生光，另一种其导致DNA转录，并通过转录和随后的翻译获得多肽。昆虫萤光素衍生自生物发光昆虫，优选双翅目和鞘翅目（萤火虫和甲虫）。类似地，通过包含编码第二多肽的另外的多脱氧核糖核苷酸使得它们各自的多肽编码阅读框形成连续的阅读框，类似地形成杂交蛋白。还公开了使用与针对抗原的抗体结合的杂交蛋白定量测定流体存在未知量的抗原的方法，测定抗体和抗原反应后的发光，并通过比较确定抗原的量的等效杂交蛋白在等同条件下在已知量的抗原的等同条件下反应的在先前测量的发光的确定的发光。

34. 发明授权

US08361739B2 Luciferase-based assays 有权
公开(公告)号：US08361739B2
公开(公告)日：2013-01-29
申请号：US12819334
申请日：2010-06-21
申请人： Erika Hawkins , James J. Cali , Samuel Kin Sang Ho , Martha A. O'Brien , Richard Somberg , Robert F. Bulleit , Keith V. Wood
发明人： Erika Hawkins , James J. Cali , Samuel Kin Sang Ho , Martha A. O'Brien , Richard Somberg , Robert F. Bulleit , Keith V. Wood
IPC分类号： C12Q1/66 , C12Q1/34 , C12Q1/37
CPC分类号： C12Q1/66
摘要： A method and kit are provided for enhancing the tolerance of an assay reagent to compounds in an assay sample, the assay reagent including a luciferase enzyme. The method includes contacting the luciferase with a tolerance enhancement agent in an amount sufficient to substantially protect luciferase enzyme activity from interference of the compound and minimize interference by at least about 10% relative to an assay not having tolerance enhancement agent.

35. 发明申请

US20120220013A1 COVALENT TETHERING OF FUNCTIONAL GROUPS TO PROTEINS AND SUBSTRATES THEREFOR 有权
标题翻译：功能组合对蛋白质和底物的共价分析
公开(公告)号：US20120220013A1
公开(公告)日：2012-08-30
申请号：US13435970
申请日：2012-03-30
申请人： Aldis Darzins , Lance Encell , Dieter Klaubert , Georgyi V. Los , Mark McDougall , Keith V. Wood , Monika G. Wood , Chad Zimprich
发明人： Aldis Darzins , Lance Encell , Dieter Klaubert , Georgyi V. Los , Mark McDougall , Keith V. Wood , Monika G. Wood , Chad Zimprich
IPC分类号： C12N9/14 , C12N15/62 , C12N15/55
CPC分类号： G01N33/581 , C07K7/06 , C07K16/00 , C07K2317/622 , C07K2319/43 , C07K2319/61 , C09B1/00 , C09B11/24 , C09B23/04 , C09B23/06 , C09B23/105 , C12N9/14 , C12Q1/34 , C12Y308/01005 , G01N33/573 , G01N2333/914 , Y10S530/825
摘要： A mutant hydrolase optionally fused to a protein of interest is provided. The mutant hydrolase is capable of forming a bond with a substrate for the corresponding nonmutant (wild-type) hydrolase which is more stable than the bond formed between the wild-type hydrolase and the substrate and has at least two amino acid substitutions relative to the wild-type hydrolase. Substrates for hydrolases comprising one or more functional groups are also provided, as well as methods of using the mutant hydrolase and the substrates of the invention. Also provided is a fusion protein capable of forming a stable bond with a substrate and cells which express the fusion protein.
摘要翻译：提供任选融合到感兴趣的蛋白质的突变体水解酶。突变体水解酶能够与相应的非突变（野生型）水解酶的底物形成键，其比野生型水解酶和底物之间形成的键更稳定，并且相对于野生型水解酶具有至少两个氨基酸取代野生型水解酶。还提供了包含一个或多个官能团的水解酶的底物，以及使用本发明的突变体水解酶和底物的方法。还提供能够与底物形成稳定结合的融合蛋白和表达融合蛋白的细胞。

36. 发明授权

US08106052B2 Luminescence-based methods and probes for measuring cytochrome P450 activity 有权
标题翻译：用于测量细胞色素P450活性的基于发光的方法和探针
公开(公告)号：US08106052B2
公开(公告)日：2012-01-31
申请号：US12754164
申请日：2010-04-05
申请人： James J. Cali , Dieter Klaubert , William Daily , Samuel Kin Sang Ho , Susan Frackman , Erika Hawkins , Keith V. Wood
发明人： James J. Cali , Dieter Klaubert , William Daily , Samuel Kin Sang Ho , Susan Frackman , Erika Hawkins , Keith V. Wood
IPC分类号： A61K31/403 , A61K31/4164 , A61K31/422 , A61K31/437 , A61K31/4985 , C07D413/04 , C07D417/04 , C07D471/04 , C07D487/04
CPC分类号： G01N33/502 , A61K49/0008 , C07D417/04 , C12Q1/66 , G01N33/5088 , G01N33/582 , G01N2333/90245 , G01N2333/90251 , G01N2500/10
摘要： The present invention provides methods, compositions, substrates, and kits useful for analyzing the metabolic activity in cells, tissue, and animals and for screening test compounds for their effect on cytochrome P450 activity. In particular, a one-step and two-step methods using luminogenic molecules, e.g. luciferin or coelenterazines, that are cytochrome P450 substrates and that are also bioluminescent enzyme, e.g., luciferase, pro-substrates are provided. Upon addition of the luciferin derivative or other luminogenic molecule into a P450 reaction, the P450 enzyme metabolizes the molecule into a bioluminescent enzyme substrate, e.g., luciferin and/or luciferin derivative metabolite, in a P450 reaction. The resulting metabolite(s) serves as a substrate of the bioluminescent enzyme, e.g., luciferase, in a second light-generating reaction. Luminescent cytochrome P450 assays with low background signals and high sensitivity are disclosed and isoform selectivity is demonstrated. The present invention also provides an improved method for performing luciferase reactions which employs added pyrophosphatase to remove inorganic pyrophosphate, a luciferase inhibitor which may be present in the reaction mixture as a contaminant or may be generated during the reaction. The present method further provides a method for stabilizing and prolonging the luminescent signal in a luciferase-based assay using luciferase stabilizing agents such as reversible luciferase inhibitors.
摘要翻译：本发明提供了可用于分析细胞，组织和动物中的代谢活性以及筛选测试化合物对细胞色素P450活性的影响的方法，组合物，底物和试剂盒。特别地，使用发光分子的一步和两步法。荧光素或coelenterazines，其是细胞色素P450底物并且也是生物发光酶，例如萤光素酶，原底物。在P450反应中加入荧光素衍生物或其他发光分子时，P450酶在P450反应中将分子代谢为生物发光酶底物，例如荧光素和/或荧光素衍生物代谢产物。所产生的代谢物在第二次发光反应中用作生物发光酶，例如荧光素酶的底物。公开了具有低背景信号和高灵敏度的发光细胞色素P450测定，证明了同工型选择性。本发明还提供一种用于进行荧光素酶反应的改进方法，其使用添加的焦磷酸酶来除去无机焦磷酸盐，荧光素酶抑制剂，其可作为污染物存在于反应混合物中，或可能在反应期间产生。本发明方法还提供了一种使用荧光素酶稳定剂如可逆荧光素酶抑制剂在基于荧光素酶的测定中稳定和延长发光信号的方法。

37. 发明授权

US07741067B2 Luciferase-based assays 有权
标题翻译：基于荧光素酶的测定
公开(公告)号：US07741067B2
公开(公告)日：2010-06-22
申请号：US10746995
申请日：2003-12-23
申请人： Erika Hawkins , James J. Cali , Samuel Kin Sang Ho , Martha A. O'Brien , Richard Somberg , Robert F. Bulleit , Keith V. Wood
发明人： Erika Hawkins , James J. Cali , Samuel Kin Sang Ho , Martha A. O'Brien , Richard Somberg , Robert F. Bulleit , Keith V. Wood
IPC分类号： C12Q1/66 , C12Q1/00 , C12Q1/08
CPC分类号： C12Q1/66
摘要： A method and kit is provided for enhancing the tolerance of an assay reagent to compounds in an assay sample, the assay reagent including a luciferase enzyme. The method includes contacting the luciferase with a tolerance enhancement agent in an amount sufficient to substantially protect luciferase enzyme activity from interference of the compound and minimize interference by at least about 10% relative to an assay not having tolerance enhancement agent.
摘要翻译：提供了一种方法和试剂盒，用于增强测定试剂对测定样品中化合物的耐受性，测定试剂包括荧光素酶。该方法包括使荧光素酶与容许增强剂接触，其量足以基本上保护荧光素酶活性免受化合物的干扰，并使相对于不具有耐受性增强剂的测定法的干扰最小化至少约10％。

38. 发明授权

US07728118B2 Synthetic nucleic acid molecule compositions and methods of preparation 有权
标题翻译：合成核酸分子组合物和制备方法
公开(公告)号：US07728118B2
公开(公告)日：2010-06-01
申请号：US10943508
申请日：2004-09-17
申请人： Keith V. Wood , Monika G. Wood , Brian Almond , Aileen Paguio , Frank Fan
发明人： Keith V. Wood , Monika G. Wood , Brian Almond , Aileen Paguio , Frank Fan
IPC分类号： C07H21/02 , C07H21/04 , C12N15/00 , C12Q1/68 , C12P19/34
CPC分类号： C07K14/43595 , C07K2319/00 , C12N15/65 , C12N15/70
摘要： A method to prepare synthetic nucleic acid molecules having reduced inappropriate or unintended transcriptional characteristics when expressed in a particular host cell.
摘要翻译：制备在特定宿主细胞中表达时具有降低的不适当或非预期转录特征的合成核酸分子的方法。

39. 发明授权

US07692022B2 Luminescence-based methods and probes for measuring cytochrome P450 activity 有权
标题翻译：用于测量细胞色素P450活性的基于发光的方法和探针
公开(公告)号：US07692022B2
公开(公告)日：2010-04-06
申请号：US10665314
申请日：2003-09-19
申请人： James J. Cali , Dieter Klaubert , William Daily , Samuel Kin Sang Ho , Susan Frackman , Erika Hawkins , Keith V. Wood
发明人： James J. Cali , Dieter Klaubert , William Daily , Samuel Kin Sang Ho , Susan Frackman , Erika Hawkins , Keith V. Wood
IPC分类号： C07D277/68 , C07D417/04
CPC分类号： G01N33/502 , A61K49/0008 , C07D417/04 , C12Q1/66 , G01N33/5088 , G01N33/582 , G01N2333/90245 , G01N2333/90251 , G01N2500/10
摘要： The present invention provides methods, compositions, substrates, and kits useful for analyzing the metabolic activity in cells, tissue, and animals and for screening test compounds for their effect on cytochrome P450 activity. In particular, a one-step and two-step methods using luminogenic molecules, e.g. luciferin or coelenterazines, that are cytochrome P450 substrates and that are also bioluminescent enzyme, e.g., luciferase, pro-substrates are provided. Upon addition of the luciferin derivative or other luminogenic molecule into a P450 reaction, the P450 enzyme metabolizes the molecule into a bioluminescent enzyme substrate, e.g., luciferin and/or luciferin derivative metabolite, in a P450 reaction. The resulting metabolite(s) serves as a substrate of the bioluminescent enzyme, e.g., luciferase, in a second light-generating reaction. Luminescent cytochrome P450 assays with low background signals and high sensitivity are disclosed and isoform selectivity is demonstrated. The present invention also provides an improved method for performing luciferase reactions which employs added pyrophosphatase to remove inorganic pyrophosphate, a luciferase inhibitor which may be present in the reaction mixture as a contaminant or may be generated during the reaction. The present method further provides a method for stabilizing and prolonging the luminescent signal in a luciferase-based assay using luciferase stabilizing agents such as reversible luciferase inhibitors.
摘要翻译：本发明提供了可用于分析细胞，组织和动物中的代谢活性以及筛选测试化合物对细胞色素P450活性的影响的方法，组合物，底物和试剂盒。特别地，使用发光分子的一步和两步法。荧光素或coelenterazines，其是细胞色素P450底物并且也是生物发光酶，例如萤光素酶，原底物。在P450反应中加入荧光素衍生物或其他发光分子时，P450酶在P450反应中将分子代谢为生物发光酶底物，例如荧光素和/或荧光素衍生物代谢产物。所产生的代谢物在第二次发光反应中用作生物发光酶，例如荧光素酶的底物。公开了具有低背景信号和高灵敏度的发光细胞色素P450测定，证明了同工型选择性。本发明还提供一种用于进行荧光素酶反应的改进方法，其使用添加的焦磷酸酶来除去无机焦磷酸盐，荧光素酶抑制剂，其可作为污染物存在于反应混合物中，或可能在反应期间产生。本发明方法还提供了一种使用荧光素酶稳定剂如可逆荧光素酶抑制剂在基于荧光素酶的测定中稳定和延长发光信号的方法。

40. 发明申请

US20090191622A1 SYNTHETIC NUCLEIC ACIDS FROM AQUATIC SPECIES 审中-公开
标题翻译：来自水生物种的合成核酸
公开(公告)号：US20090191622A1
公开(公告)日：2009-07-30
申请号：US12323270
申请日：2008-11-25
申请人： Brian D. Almond , Monika G. Wood , Keith V. Wood
发明人： Brian D. Almond , Monika G. Wood , Keith V. Wood
IPC分类号： C12N15/00 , C12N15/11 , C07H1/00
CPC分类号： C12N9/0069 , C07K14/43595 , C12N15/67 , C12N15/8212 , C12N15/8216
摘要： A synthetic nucleic acid molecule is provided that includes nucleotides of a coding region for a fluorescent polypeptide having a codon composition differing at more than 25% of the codons from a parent nucleic acid sequence encoding a fluorescent polypeptide. The synthetic nucleic acid molecule has at least 3-fold fewer transcription regulatory sequences relative to the average number of such sequences in the parent nucleic acid sequence. The polypeptide encoded by the synthetic nucleic acid molecule preferably has at least 85% sequence identity to the polypeptide encoded by the parent nucleic acid sequence.
摘要翻译：提供了合成核酸分子，其包括编码区域的核苷酸，所述核苷酸的密码子组成不同于编码荧光多肽的母体核酸序列的密码子的25％以上的荧光多肽。合成的核酸分子相对于母核酸序列中这样的序列的平均数目具有至少3倍的转录调节序列。由合成核酸分子编码的多肽优选与由母体核酸序列编码的多肽具有至少85％的序列同一性。

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