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    Compositions and methods for detection and isolation of phosphorylated molecules
    1.
    发明申请
    Compositions and methods for detection and isolation of phosphorylated molecules 审中-公开
    用于磷酸化分子检测和分离的组合物和方法

    公开(公告)号：US20100009381A1

    公开(公告)日：2010-01-14

    申请号：US12500854

    申请日：2009-07-10

    申请人： Brian Agnew ,  Joseph Beechem ,  Kyle Gee ,  Richard Haugland ,  Thomas Steinberg ,  Wayne Patton

    发明人： Brian Agnew ,  Joseph Beechem ,  Kyle Gee ,  Richard Haugland ,  Thomas Steinberg ,  Wayne Patton

    IPC分类号： G01N33/53

    CPC分类号： G01N33/5008 ,  C12Q1/42 ,  C12Q1/485 ,  G01N33/502 ,  G01N33/5091 ,  G01N2333/4709 ,  Y10T436/142222 ,  Y10T436/145555 ,  Y10T436/147777 ,  Y10T436/201666

    摘要： The present invention relates to phosphate-binding compounds that find use in binding, detecting and isolating phosphorylated target molecules including the subsequent identification of target molecules that interact with phosphorylated target molecules or molecules capable of being phosphorylated. A binding solution is provide that comprises a phosphate-binding compound, an acid and a metal ion wherein the metal ion simultaneously interacts with an exposed phosphate group on a target molecule and the metal chelating moiety of the phosphate-binding compound forming a bridge between the phosphate-binding compound and a phosphorylated target molecule resulting in a ternary complex. The binding solution of the present invention finds use in binding and detecting immobilized and solubilized phosphorylated target molecules, isolation of phosphorylated target molecules from a complex mixture and aiding in proteomic analysis wherein kinase and phosphatase substrates and enzymes can be identified.

    摘要翻译： 本发明涉及用于结合，检测和分离磷酸化靶分子的磷酸盐结合化合物，包括随后鉴定与磷酸化靶分子或能够被磷酸化的分子相互作用的靶分子。 提供的结合溶液包括磷酸盐结合化合物，酸和金属离子，其中金属离子同时与靶分子上暴露的磷酸基团相互作用，并且磷酸盐结合化合物的金属螯合部分形成在 磷酸化结合化合物和导致三元复合物的磷酸化靶分子。 本发明的结合溶液用于结合和检测固定和溶解的磷酸化靶分子，从复杂混合物中分离磷酸化靶分子，并辅助蛋白质组分析，其中可以鉴定激酶和磷酸酶底物和酶。

    Competitive immunoassay
    2.
    发明申请
    Competitive immunoassay 有权
    竞争性免疫测定

    公开(公告)号：US20060160068A1

    公开(公告)日：2006-07-20

    申请号：US10943463

    申请日：2004-09-17

    申请人： Joseph Beechem ,  Kyle Gee ,  David Hagen ,  Iain Johnson ,  Hee-Chol Kang ,  Christina Pastula

    发明人： Joseph Beechem ,  Kyle Gee ,  David Hagen ,  Iain Johnson ,  Hee-Chol Kang ,  Christina Pastula

    IPC分类号： C12Q1/70 ,  C12Q1/68 ,  C07K16/46 ,  C07F5/02 ,  C07D311/88

    CPC分类号： G01N33/542 ,  C07F9/12 ,  C07F9/65318 ,  C07F9/65522 ,  C07F9/6596 ,  C07K16/46 ,  Y10S435/968 ,  Y10S436/80 ,  Y10S530/807 ,  Y10T436/13

    摘要： The present invention provides ligand-detection reagents, ligand analogs and methods for determining the presence of a ligand in a sample. The ligand-detection reagent comprises a ligand-binding antibody and a ligand analog to form an antibody-ligand analog complex wherein the ligand analog is covalently bonded to a reporter molecule. This complex may additionally comprise a labeling protein non-covalently bonded to the antibody to form a ternary complex wherein the labeling protein comprises a monovalent antibody fragment or a non-antibody protein that is covalently bonded to a label moiety. The reporter molecule is either quenched by the ligand-binding antibody or by the label moiety of the labeling protein, depending on the reporter molecule and the ligand-binding antibody, wherein the amount of quenching is directly related to the amount of ligand present in the sample. Alternatively, the ligand analog is fluorogenic wherein the ligand analog is essentially non-fluorescent in solution but when bound by the ligand-binding antibody the detectable signal increases. In this instance a decrease in signal, as opposed to the relieving of quenching, is measured for the presence of a target ligand.

    摘要翻译： 本发明提供配体检测试剂，配体类似物和用于测定样品中配体存在的方法。 配体检测试剂包含配体结合抗体和配体类似物以形成抗体 - 配体类似物复合物，其中配体类似物共价键合到报告分子。 该复合物还可以包含与抗体非共价键合以形成三元复合物的标记蛋白质，其中标记蛋白质包含共价键合到标记部分的单价抗体片段或非抗体蛋白质。 取决于报道分子和配体结合抗体，报告分子被配体结合抗体或标记蛋白的标记部分淬灭，其中淬灭量与存在于 样品。 或者，配体类似物是荧光的，其中配体类似物在溶液中基本上是非荧光的，但当被配体结合抗体结合时，可检测信号增加。 在这种情况下，与目标配体的存在相比，测量了与消除淬灭相反的信号降低。

    ISOLATED PHOSPHOLIPID-PROTEIN PARTICLES
    3.
    发明申请
    ISOLATED PHOSPHOLIPID-PROTEIN PARTICLES 审中-公开
    分离的磷脂 - 蛋白质颗粒

    公开(公告)号：US20090161828A1

    公开(公告)日：2009-06-25

    申请号：US12333191

    申请日：2008-12-11

    申请人： Federico Katzen ,  Julia Fletcher ,  Wieslaw Kudlicki ,  Joseph Beechem ,  Lilin Wang

    发明人： Federico Katzen ,  Julia Fletcher ,  Wieslaw Kudlicki ,  Joseph Beechem ,  Lilin Wang

    IPC分类号： G01N23/207 ,  C07K14/00 ,  C12N9/10 ,  C12N9/48 ,  C12N9/16

    CPC分类号： C12P21/02 ,  C07K1/02

    摘要： Systems and methods are provided for producing a protein of interest that is typically not amenable to expression in soluble form in in vitro expression systems. In some aspects, the invention provides methods of synthesizing proteins using in vitro protein synthesis systems that include a scaffold protein such as apolipoprotein or an amphipathic alpha helix containing (“AAHC”) protein, in which higher yields of soluble protein are produced than in the absence of the scaffold protein. The scaffold proteins may be provided in an in vitro protein synthesis system associated with lipid or not associated with lipid. The scaffold protein may be provided as a protein per se or may be encoded by a nucleic acid template and co-expressed with the protein of interest. The invention also provides compositions and kits for synthesis of proteins in soluble form, in which the compositions and kits include cell extracts for protein expression and isolation.

    摘要翻译： 提供的系统和方法用于产生通常不适于在体外表达系统中以可溶形式表达的目的蛋白质。 在一些方面，本发明提供了使用体外蛋白质合成系统合成蛋白质的方法，所述体外蛋白质合成系统包括诸如载脂蛋白或含有（“AAHC”）蛋白质的两亲性α螺旋的支架蛋白，其中产生比在 缺少支架蛋白。 支架蛋白质可以提供在与脂质相关的体外蛋白质合成系统中或与脂质不相关。 支架蛋白质可以作为蛋白质本身提供，或者可以由核酸模板编码并与感兴趣的蛋白质共表达。 本发明还提供用于合成可溶形式的蛋白质的组合物和试剂盒，其中组合物和试剂盒包括用于蛋白质表达和分离的细胞提取物。

    Antibody complexes and methods for immunolabeling
    4.
    发明申请
    Antibody complexes and methods for immunolabeling 审中-公开
    抗体复合物和免疫标记方法

    公开(公告)号：US20070269902A1

    公开(公告)日：2007-11-22

    申请号：US10666291

    申请日：2003-09-17

    申请人： Joseph Beechem ,  David Hagen ,  Iain Johnson

    发明人： Joseph Beechem ,  David Hagen ,  Iain Johnson

    IPC分类号： G01N33/00 ,  C12Q1/68

    CPC分类号： B82Y30/00 ,  B82Y5/00 ,  B82Y10/00 ,  G01N33/53 ,  G01N33/58 ,  G01N33/6857

    摘要： The present invention provides labeling reagents and methods for labeling primary antibodies and for detecting a target in a sample using an immuno-labeled complex that comprises a target-binding antibody and one or more labeling reagents. The labeling reagents comprise monovalent antibody fragments or non-antibody monomeric proteins whereby the labeling reagents have affinity for a specific region of the target-binding antibody and are covalently attached to a label. Typically, the labeling reagent is an anti-Fc Fab or Fab′ fragment that was generated by immunizing a goat or rabbit with the Fc fragment of an antibody. The present invention provides for discrete subsets of labeling reagent and immuno-labeled complexes that facilitate the simultaneous detection of multiple targets in a sample wherein the immuno-labeled complexes are distinguished by i) a ratio of label to labeling reagent, or ii) a physical property of said label, or iii) a ratio of labeling reagent to said target-binding antibody, or iv) by said target-binding antibody. This is particularly useful for fluorophore labels that can be attached to labeling reagents and subsequently immuno-labeled complexes in ratios for the detection of multiple targets.

    摘要翻译： 本发明提供标记试剂和用于标记一抗的标记试剂和使用包含靶结合抗体和一种或多种标记试剂的免疫标记复合物检测样品中靶标的方法。 标记试剂包含单价抗体片段或非抗体单体蛋白质，由此标记试剂对靶结合抗体的特定区域具有亲和力，并且共价连接到标记物上。 通常，标记试剂是通过用抗体的Fc片段免疫山羊或兔而产生的抗Fc Fab或Fab'片段。 本发明提供标记试剂和免疫标记的复合物的离散子集，其有助于同时检测样品中的多个靶标，其中通过i）标记物与标记试剂的比率来区分免疫标记的复合物，或ii）物理 或iii）标记试剂与所述靶结合抗体的比例，或iv）所述靶结合抗体。 这对于可以连接到标记试剂和随后的用于检测多个靶的比例的免疫标记的复合物的荧光团标记物特别有用。

    Methods and apparatus for single molecule sequencing using energy transfer detection
    5.
    发明授权
    Methods and apparatus for single molecule sequencing using energy transfer detection 有权
    使用能量转移检测的单分子测序的方法和装置

    公开(公告)号：US08999674B2

    公开(公告)日：2015-04-07

    申请号：US13562159

    申请日：2012-07-30

    申请人： Joseph Beechem ,  Theo Nikiforov ,  Vi-En Choong ,  Xinzhan Peng ,  Guobin Luo ,  Cheng-Yao Chen ,  Michael Previte

    发明人： Joseph Beechem ,  Theo Nikiforov ,  Vi-En Choong ,  Xinzhan Peng ,  Guobin Luo ,  Cheng-Yao Chen ,  Michael Previte

    IPC分类号： C12P19/34 ,  C12N9/12 ,  C07H19/20

    CPC分类号： C12Q1/6869 ,  C07H19/20 ,  C12N9/1241 ,  C12N9/1252 ,  C12N9/96 ,  C12Q1/6818 ,  C12Y207/07 ,  C12Y207/07007 ,  G01N21/6428 ,  G01N33/582 ,  G01N2021/6432

    摘要： Provided herein are systems and methods for nucleotide incorporation reactions. The systems comprise polymerases having altered nucleotide incorporation kinetics and are linked to an energy transfer donor moiety, and nucleotide molecules linked with at least one energy transfer acceptor moiety. The donor and acceptor moieties undergo energy transfer when the polymerase and nucleotide are proximal to each other during nucleotide binding and/or nucleotide incorporation. As the donor and acceptor moieties undergo energy transfer, they generate an energy transfer signal which can be associated with nucleotide binding or incorporation. Detecting a time sequence of the generated signals, or the change in the signals, can be used to determine the order of the incorporated nucleotides, and can therefore be used to deduce the sequence of the target molecule.

    摘要翻译： 本文提供了用于核苷酸掺入反应的系统和方法。 该系统包含具有改变的核苷酸掺入动力学并且与能量转移供体部分连接的聚合酶，以及与至少一个能量转移受体部分连接的核苷酸分子。 当核苷酸结合和/或核苷酸掺入期间聚合酶和核苷酸彼此接近时，供体和受体部分进行能量转移。 当供体和受体部分进行能量转移时，它们产生可与核苷酸结合或掺入相关联的能量转移信号。 检测所产生的信号的时间序列或信号的变化可用于确定掺入的核苷酸的顺序，因此可用于推导靶分子的序列。

    ISOLATED PHOSPHOLIPID-PROTEIN PARTICLES
    7.
    发明申请
    ISOLATED PHOSPHOLIPID-PROTEIN PARTICLES 审中-公开
    分离的磷脂 - 蛋白质颗粒

    公开(公告)号：US20110104781A1

    公开(公告)日：2011-05-05

    申请号：US12890386

    申请日：2010-09-24

    申请人： FEDERICO KATZEN ,  JULIA FLETCHER ,  WIESLAW KUDLICKI ,  JOSEPH BEECHEM ,  LILIN WANG

    发明人： FEDERICO KATZEN ,  JULIA FLETCHER ,  WIESLAW KUDLICKI ,  JOSEPH BEECHEM ,  LILIN WANG

    IPC分类号： C12N9/00 ,  C07K14/00 ,  C12N5/071 ,  C12N1/16

    CPC分类号： C12P21/02 ,  C07K1/02

    摘要： Systems and methods are provided for producing a protein of interest that is typically not amenable to expression in soluble form in in vitro expression systems. In some aspects, the invention provides methods of synthesizing proteins using in vitro protein synthesis systems that include a scaffold protein such as apolipoprotein or an amphipathic alpha helix containing (“AAHC”) protein, in which higher yields of soluble protein are produced than in the absence of the scaffold protein. The scaffold proteins may be provided in an in vitro protein synthesis system associated with lipid or not associated with lipid. The scaffold protein may be provided as a protein per se or may be encoded by a nucleic acid template and co-expressed with the protein of interest. The invention also provides compositions and kits for synthesis of proteins in soluble form, in which the compositions and kits include cell extracts for protein expression and isolation.

    摘要翻译： 提供的系统和方法用于产生通常不适于在体外表达系统中以可溶形式表达的目的蛋白质。 在一些方面，本发明提供了使用体外蛋白质合成系统合成蛋白质的方法，所述体外蛋白质合成系统包括诸如载脂蛋白或含有（“AAHC”）蛋白质的两亲性α螺旋的支架蛋白，其中产生比在 缺少支架蛋白。 支架蛋白质可以提供在与脂质相关的体外蛋白质合成系统中或与脂质不相关。 支架蛋白质可以作为蛋白质本身提供，或者可以由核酸模板编码并与感兴趣的蛋白质共表达。 本发明还提供用于合成可溶形式的蛋白质的组合物和试剂盒，其中组合物和试剂盒包括用于蛋白质表达和分离的细胞提取物。

    METHODS AND APPARATUS FOR SINGLE MOLECULE SEQUENCING USING ENERGY TRANSFER DETECTION
    9.
    发明申请
    METHODS AND APPARATUS FOR SINGLE MOLECULE SEQUENCING USING ENERGY TRANSFER DETECTION 审中-公开
    使用能量转移检测的单分子序列的方法和装置

    公开(公告)号：US20100255487A1

    公开(公告)日：2010-10-07

    申请号：US12748168

    申请日：2010-03-26

    申请人： JOSEPH BEECHEM ,  THEO NIKIFOROV ,  Vi-En Choong ,  Xinzhan Peng ,  Guobin Luo ,  Cheng-Yao Chen ,  Michael Previte

    发明人： JOSEPH BEECHEM ,  THEO NIKIFOROV ,  Vi-En Choong ,  Xinzhan Peng ,  Guobin Luo ,  Cheng-Yao Chen ,  Michael Previte

    IPC分类号： C12Q1/68 ,  C12N9/10

    CPC分类号： C12Q1/6869 ,  C07H19/20 ,  C12N9/1241 ,  C12N9/1252 ,  C12N9/96 ,  C12Q1/6818 ,  C12Y207/07 ,  C12Y207/07007 ,  G01N21/6428 ,  G01N33/582 ,  G01N2021/6432

    摘要： Provided herein are systems and methods for nucleotide incorporation reactions. The systems comprise polymerases having altered nucleotide incorporation kinetics and are linked to an energy transfer donor moiety, and nucleotide molecules linked with at least one energy transfer acceptor moiety. The donor and acceptor moieties undergo energy transfer when the polymerase and nucleotide are proximal to each other during nucleotide binding and/or nucleotide incorporation. As the donor and acceptor moieties undergo energy transfer, they generate an energy transfer signal which can be associated with nucleotide binding or incorporation. Detecting a time sequence of the generated signals, or the change in the signals, can be used to determine the order of the incorporated nucleotides, and can therefore be used to deduce the sequence of the target molecule.

    摘要翻译： 本文提供了用于核苷酸掺入反应的系统和方法。 该系统包含具有改变的核苷酸掺入动力学并且与能量转移供体部分连接的聚合酶，以及与至少一个能量转移受体部分连接的核苷酸分子。 当核苷酸结合和/或核苷酸掺入期间聚合酶和核苷酸彼此接近时，供体和受体部分进行能量转移。 当供体和受体部分进行能量转移时，它们产生可与核苷酸结合或掺入相关联的能量转移信号。 检测所产生的信号的时间序列或信号的变化可用于确定掺入的核苷酸的顺序，因此可用于推导靶分子的序列。