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1. 发明申请

US20100009381A1 Compositions and methods for detection and isolation of phosphorylated molecules 审中-公开
标题翻译：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。

2. 发明申请

US20060160068A1 Competitive immunoassay 有权
标题翻译：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。

3. 发明申请

US20090161828A1 ISOLATED PHOSPHOLIPID-PROTEIN PARTICLES 审中-公开
标题翻译：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。

4. 发明申请

US20070269902A1 Antibody complexes and methods for immunolabeling 审中-公开
标题翻译：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。

5. 发明授权

US08603792B2 Conjugates of biomolecules to nanoparticles 有权
标题翻译：生物分子与纳米颗粒的结合
公开(公告)号：US08603792B2
公开(公告)日：2013-12-10
申请号：US12748355
申请日：2010-03-26
申请人： Theo Nikiforov , Daniel Mazur , Xinzhan Peng , Tommie Lloyd Lincecum, Jr. , Yuri Belosludtsev , Howard Reese , Dmitriy Gremyachinskiy , Roman Rozhkov , John M. Mauro , Joseph Beechem , Eric Tulsky , Imad Naasani , Kari Haley , Joseph A. Treadway
发明人： Theo Nikiforov , Daniel Mazur , Xinzhan Peng , Tommie Lloyd Lincecum, Jr. , Yuri Belosludtsev , Howard Reese , Dmitriy Gremyachinskiy , Roman Rozhkov , John M. Mauro , Joseph Beechem , Eric Tulsky , Imad Naasani , Kari Haley , Joseph A. Treadway
IPC分类号： C12N9/12
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： Disclosed herein are conjugates comprising a biomolecule linked to a label that have biological activity and are useful in a wide variety of biological applications. For example, provided herein are polymerase-nanoparticle conjugates including a polymerase linked to a nanoparticle, wherein the conjugate has polymerase activity. Such conjugates can exhibit reduced aggregation and improved stochiometries wherein the average biomolecule:nanoparticle ratio approaches or equals 1:1. Also disclosed herein are improved methods for preparing such conjugates, and methods and systems for using such conjugates in biological applications such as nucleotide incorporation, primer extension and single molecule sequencing.
摘要翻译：本文公开了包含连接到具有生物活性并可用于多种生物应用的标签的生物分子的缀合物。例如，本文提供了聚合酶 - 纳米颗粒缀合物，其包括与纳米颗粒连接的聚合酶，其中缀合物具有聚合酶活性。这样的缀合物可以表现出减少的聚集和改进的沉淀物，其中平均生物分子：纳米颗粒比接近或等于1:1。本文还公开了制备这种缀合物的改进方法，以及在生物应用中使用这种缀合物的方法和系统，例如核苷酸掺入，引物延伸和单分子测序。

6. 发明授权

US08535894B2 Antibody complexes and methods for immunolabeling 有权
标题翻译：抗体复合物和免疫标记方法
公开(公告)号：US08535894B2
公开(公告)日：2013-09-17
申请号：US13610009
申请日：2012-09-11
申请人： Robert Archer , Richard Haugland , Rosaria Haugland , Joseph Beechem , David Hagen
发明人： Robert Archer , Richard Haugland , Rosaria Haugland , Joseph Beechem , David Hagen
IPC分类号： G01N33/53
CPC分类号： B82Y5/00 , B82Y10/00 , G01N33/53
摘要： Provided are 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 proteins have affinity for a specific region of the target-binding antibody and are covalently attached to a label. Discrete subsets of labeling reagent and immune-labeled complexes are provided that facilitate the simultaneous detection of multiple targets in a sample-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：提供标记试剂和用于标记一抗的标记试剂和用于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的靶标的方法。标记试剂包括单价抗体片段或非抗体单体蛋白质，由此标记的蛋白质对靶标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。提供标记试剂和免疫标记复合物的离散子集，其有利于同时检测样品中多个靶标的复合物，其特征在于i）标记与标记试剂的比例，或ii）所述标记的物理性质，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。

7. 发明申请

US20120329042A1 METHODS AND APPARATUS FOR SINGLE MOLECULE SEQUENCING USING ENERGY TRANSFER DETECTION 有权
标题翻译：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：US20120329042A1
公开(公告)日：2012-12-27
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： G01N21/64
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。

8. 发明申请

US20110311963A1 Method and Apparatus for Addressable Flow Cells in Single Molecule Sequencing 审中-公开
标题翻译：单分子测序中可寻址流动池的方法和装置
公开(公告)号：US20110311963A1
公开(公告)日：2011-12-22
申请号：US13049660
申请日：2011-03-16
申请人： W. MICHAEL LAFFERTY , JOSEPH BEECHEM , HONGYE SUN , MICHAEL METZKER
发明人： W. MICHAEL LAFFERTY , JOSEPH BEECHEM , HONGYE SUN , MICHAEL METZKER
IPC分类号： C12Q1/68
CPC分类号： C12Q1/6874 , C12Q2523/319 , C12Q2525/186 , C12Q2565/513
摘要： A method of sequencing a plurality of template nucleotide sequences includes immobilizing the plurality of template nucleotide sequences on a substrate. A first subset of the plurality of template nucleotide sequences is immobilized in a first field of view and a second subset of the plurality of template nucleotide sequences is immobilized in a second field of view. The first and second subsets are hybridized to a caged primer. The caged primer includes a caging group. The method further includes lysing the caging group from the caged primer in the first field of view and observing the first field of view to detect sequencing of the first subset of the plurality of template nucleotide sequences.
摘要翻译：对多个模板核苷酸序列进行测序的方法包括将多个模板核苷酸序列固定在底物上。多个模板核苷酸序列的第一子集被固定在第一视场中，并且多个模板核苷酸序列的第二子集被固定在第二视野中。第一和第二子集与笼式引物杂交。笼状引物包括笼养组。该方法还包括在第一视场中从笼式引物裂解笼基组，并观察第一视野以检测多个模板核苷酸序列的第一子集的测序。

9. 发明申请

US20110236983A1 SINGLE MOLECULE DETECTION AND SEQUENCING USING FLUORESCENCE LIFETIME IMAGING 审中-公开
标题翻译：使用荧光生物成像的单分子检测和测序
公开(公告)号：US20110236983A1
公开(公告)日：2011-09-29
申请号：US12980817
申请日：2010-12-29
申请人： Joseph Beechem , Theofilos Kotseroglou , William Michael Lafferty , Mark F. Oldham
发明人： Joseph Beechem , Theofilos Kotseroglou , William Michael Lafferty , Mark F. Oldham
IPC分类号： G01N33/50 , G01N21/64 , B82Y99/00
CPC分类号： G01N21/6408 , C12Q1/6818 , G01N21/6428 , G01N21/6458 , G01N2021/6441 , Y10T436/143333 , C12Q2563/173 , C12Q2563/155 , C12Q2561/12
摘要： A nucleic acid detection system and method are provided, in which excitation energy is transmitted from a pulsed excitation source to a reaction site including a fluorescence resonance energy transfer (FRET)-based dye system to generate a fluorescent signal at the reaction site, the fluorescent signal is detected by a detector from the reaction site, and detection of the fluorescent signal is respectively blocked and permitted at the detector by a detector gate this is timed based on an emission start time of the transmitted excitation energy.
摘要翻译：提供了一种核酸检测系统和方法，其中激发能量从脉冲激发源传输到包括基于荧光共振能量转移（FRET）的染料系统的反应位点，以在反应位点产生荧光信号，荧光信号被来自反应部位的检测器检测，并且荧光信号的检测分别被检测器门阻断并在检测器处允许，其基于发射的激发能量的发射开始时间来定时。

10. 发明申请

US20110200989A1 SINGLE MOLECULE NUCLEIC ACID SEQUENCING USING MULTIPHOTON FLUORESCENCE EXCITATION 审中-公开
标题翻译：使用多光子荧光激发的单分子核酸序列
公开(公告)号：US20110200989A1
公开(公告)日：2011-08-18
申请号：US13009442
申请日：2011-01-19
申请人： Gordon A. Janaway , Christina E. Inman , Joseph Beechem
发明人： Gordon A. Janaway , Christina E. Inman , Joseph Beechem
IPC分类号： C12Q1/68 , C12M1/34 , G01N21/75 , G01N21/63 , G01N21/64
CPC分类号： G01N21/6452 , C12Q1/6818 , C12Q1/6869 , G01N21/6428 , G01N2021/6441 , Y10T436/143333 , C12Q2563/131 , C12Q2523/313
摘要： A system for detection of nucleic acids can include an excitation source configured to transmit excitation energy to a reaction site including a single molecule of nucleic acid reacted with a two-photon absorption moiety. The system also can include an optical system configured to focus the excitation energy transmitted from the excitation source to a focal region containing the reaction site, wherein said excitation energy within the focal region is sufficient to cause two-photon absorption by the two-photon absorption moiety. The system can further include a detector configured to detect emissions generated at the reaction site resulting from two-photon absorption of the excitation energy by the two-photon absorption moiety.
摘要翻译：用于检测核酸的系统可以包括被配置为将激发能量传递到包括与双光子吸收部分反应的单分子核酸的反应位点的激发源。该系统还可以包括配置成将从激发源传输的激发能量聚焦到包含反应位点的聚焦区域的光学系统，其中聚焦区域内的所述激发能足以通过双光子吸收引起双光子吸收部分。该系统还可以包括检测器，其被配置为检测由双光子吸收部分的激发能量的双光子吸收引起的在反应位置产生的发射。

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