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公开(公告)号：US10465235B2
公开(公告)日：2019-11-05
申请号：US14119925
申请日：2012-05-23
申请人： Mats Gullberg , Ola Söderberg , Ulf Landegren , Yanling Liu
发明人： Mats Gullberg , Ola Söderberg , Ulf Landegren , Yanling Liu
IPC分类号： C12Q1/68 , C12Q1/6837 , C12Q1/6804 , C12Q1/682
摘要： The present invention provides a method for detecting interactions between or with any two of at least three target substrates, or any two of at least three features of a target substrate, or a combination of interactions and features of target substrates, by a multiplexed proximity ligation assay, said method comprising: a) for each of the at least three target substrates or features, providing a proximity probe comprising a binding moiety with affinity for the feature or binding site on said substrate, and a proximity probe oligonucleotide coupled on the binding moiety; wherein each of the proximity probe oligonucleotide carries a unique tag sequence; b) mixing the proximity probes with a sample, under a condition to allow binding of each proximity probe to its respective binding site or feature on each of said substrates through the binding moiety, c) simultaneous with, or following step b), forming circularized DNA molecules where any two proximity probes bind sufficiently close to each other on the substrate, wherein each of the circularized DNA molecules comprise complementary sequences to the unique tag sequences from the two proximity probes oligo-nucleotides; d) amplifying the circularized DNA; and e) characterizing the amplified DNA.
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公开(公告)号：US20140170654A1
公开(公告)日：2014-06-19
申请号：US14116706
申请日：2012-05-11
IPC分类号： C12Q1/68
CPC分类号： C12Q1/6876 , C12Q1/6816 , C12Q1/6841 , G01N33/542 , C12Q2521/501 , C12Q2521/531 , C12Q2525/301 , C12Q2525/307 , C12Q2531/125 , C12Q2537/162
摘要： The present invention relates to a proximity-probe based detection assay for detecting an analyte in a sample and in particular to a method that comprises the use of at least one set of at least first and second proximity probes, which probes each comprise an analyte-binding domain and a nucleic acid domain and can simultaneously bind to the analyte directly or indirectly, wherein the nucleic acid domain of at least one of said proximity probes comprises a hairpin structure that can be unfolded by cleavage of the nucleic acid domain to generate at least one ligatable free end or region of complementarity to another nucleic acid molecule in said sample, wherein when the probes bind to said analyte unfolding said hairpin structure allows the nucleic acid domains of said at least first and second proximity probes to interact directly or indirectly.
摘要翻译： 本发明涉及一种用于检测样品中分析物的基于接近探针的检测测定法，特别涉及一种包括使用至少一组至少第一和第二接近探针的方法，该探针各自包含分析物 - 结合结构域和核酸结构域，并且可以直接或间接地同时结合分析物，其中至少一个所述邻近探针的核酸结构域包含发夹结构，所述发夹结构可以通过切割核酸结构域而被解折叠以产生至少 一个可连接的自由端或与所述样品中的另一个核酸分子互补的区域，其中当所述探针结合到所述分析物上展开所述发夹结构时，允许所述至少第一和第二接近探针的核酸结构域直接或间接相互作用。
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公开(公告)号：US10669569B2
公开(公告)日：2020-06-02
申请号：US13879038
申请日：2011-10-14
IPC分类号： C12Q1/68 , C12Q1/6804 , G01N33/542
摘要： Methods for detecting and quantifying an analyte employ a pair of proximity probes, each comprising a proteinaceous target-binding domain coupled to a nucleic acid domain (NAD), which NADs interact when the proximity probes have bound in proximity to their respective target; and a set of markers, wherein each marker is a nucleic acid molecule comprising a binding domain and a reporter domain giving a detectable signal, can interact with said NADs to form a nucleic acid molecule from which a detectable signal is generated, or with a nucleic acid molecule generated by interaction of said NADs, cannot interact with said NADs simultaneously with another marker in the set, generates a signal that is distinguishable from another marker signal, and is present in an amount capable of detecting analyte at a range of concentrations differing from the range of concentrations detectable by other markers.
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公开(公告)号：US20140194311A1
公开(公告)日：2014-07-10
申请号：US14119925
申请日：2012-05-23
申请人： Mats Gullberg , Ola Söderberg , Ulf Landegren , Yangling Liu
发明人： Mats Gullberg , Ola Söderberg , Ulf Landegren , Yangling Liu
IPC分类号： C12Q1/68
CPC分类号： C12Q1/6837 , C12Q1/6804 , C12Q1/682 , C12Q2531/125 , C12Q2533/107 , C12Q2537/143 , C12Q2563/179 , C12Q2565/102
摘要： The present invention provides a method for detecting interactions between or with any two of at least three target substrates, or any two of at least three features of a target substrate, or a combination of interactions and features of target substrates, by a multiplexed proximity ligation assay, said method comprising: a) for each of the at least three target substrates or features, providing a proximity probe comprising a binding moiety with affinity for the feature or binding site on said substrate, and a proximity probe oligonucleotide coupled on the binding moiety; wherein each of the proximity probe oligonucleotide carries a unique tag sequence; b) mixing the proximity probes with a sample, under a condition to allow binding of each proximity probe to its respective binding site or feature on each of said substrates through the binding moiety, c) simultaneous with, or following step b), forming circularized DNA molecules where any two proximity probes bind sufficiently close to each other on the substrate, wherein each of the circularized DNA molecules comprise complementary sequences to the unique tag sequences from the two proximity probes oligo-nucleotides; d) amplifying the circularized DNA; and e) characterizing the amplified DNA.
摘要翻译： 本发明提供了一种用于通过多路复用接近连接来检测至少三个目标衬底中的任何两个之间或与目标衬底的至少三个特征中的任何两个或目标衬底的相互作用和特征的组合之间的相互作用的方法 所述方法包括：a）对于至少三种靶基质或特征中的每一种，提供包含对所述底物上的特征或结合位点具有亲和力的结合部分的邻近探针，以及偶联在结合部分上的邻近探针寡核苷酸 ; 其中每个邻近探针寡核苷酸携带唯一的标签序列; b）将接近探针与样品混合，条件是允许通过结合部分将每个邻近探针结合到每个所述底物上的各自的结合位点或特征，c）与步骤b）同时或在步骤b）之后，形成环化的 其中任何两个接近探针在底物上彼此足够接近的DNA分子，其中每个环化的DNA分子包含来自两个邻近探针寡核苷酸的唯一标签序列的互补序列; d）扩增环化的DNA; 和e）表征扩增的DNA。
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公开(公告)号：US20130288249A1
公开(公告)日：2013-10-31
申请号：US13879038
申请日：2011-10-14
IPC分类号： C12Q1/68
CPC分类号： C12Q1/68 , C12Q1/6804 , G01N33/542 , C12Q2531/125 , C12Q2527/143 , C12Q2525/307 , C12Q2533/107
摘要： The present invention relates to methods for detecting and quantifying an analyte in a sample, principally in proximity probe assays, and in particular to an improvement in such methods to extend the dynamic range of detection, which is particularly advantageous for the detection and quantification of an analyte where the concentration range of the analyte in said sample is unknown and/or the range is likely to be broad, said method comprising: (i) contacting said sample with at least a pair of proximity probes each comprising a proteinaceous target-binding domain coupled to a nucleic acid domain such that said nucleic acid domains may be allowed to interact directly or indirectly when the proximity probes have bound in proximity to their respective target, said target being either the analyte or a binding partner for the analyte; (ii) further contacting said sample with at least one set of markers which function to extend the dynamic range of detection of the method, wherein said set comprises at least two markers, wherein each marker is a nucleic acid molecule comprising a binding domain and a reporter domain which gives rise to a detectable signal, and each marker: (a) is capable of interacting either with said nucleic acid domains to form a nucleic acid molecule from which a detectable signal is generated, or with a nucleic acid molecule generated by interaction of said domains; (b) cannot interact with said nucleic acid domains simultaneously with another marker in the set; (c) generates a signal that is distinguishable from the signal of another marker in the set; and (d) is present in an amount capable of detecting the analyte at a range of concentrations that differs from the range of concentrations detectable by other markers; (ii) allowing said markers to interact with said nucleic acid domains or said generated nucleic acid molecule; and (iii) detecting said signal.
摘要翻译： 本发明涉及用于检测和定量样品中的分析物的方法，主要是在接近探针测定中，特别是涉及扩展检测动态范围的方法的改进，这对检测和定量检测是特别有利的 分析物，其中所述样品中的分析物的浓度范围是未知的和/或该范围可能是宽的，所述方法包括：（i）使所述样品与至少一对邻近探针接触，所述至少一对接近探针各自包含蛋白质靶结合结构域 偶联到核酸结构域，使得当接近探针已经在其相应靶标附近结合时，所述核酸结构域可以被直接或间接相互作用，所述靶是分析物或分析物的结合配偶体; （ii）进一步使所述样品与至少一组标志物接触，所述标记物用于扩展所述方法的动态检测范围，其中所述组包含至少两个标记，其中每个标记是包含结合结构域和 报告结构域，其产生可检测信号，并且每个标记：（a）能够与所述核酸结构域相互作用以形成从其产生可检测信号的核酸分子，或与通过相互作用产生的核酸分子 的所述域; （b）不能与该组中的另一个标记同时与所述核酸结构域相互作用; （c）产生与该组中另一个标记的信号不同的信号; 和（d）以能够在与其他标记可检测的浓度范围不同的浓度范围内检测分析物的量存在; （ii）允许所述标记物与所述核酸结构域或所述产生的核酸分子相互作用; 和（iii）检测所述信号。
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公开(公告)号：US09551032B2
公开(公告)日：2017-01-24
申请号：US14116706
申请日：2012-05-11
IPC分类号： C12Q1/68 , A61K39/00 , C07K16/00 , G01N33/542
CPC分类号： C12Q1/6876 , C12Q1/6816 , C12Q1/6841 , G01N33/542 , C12Q2521/501 , C12Q2521/531 , C12Q2525/301 , C12Q2525/307 , C12Q2531/125 , C12Q2537/162
摘要： The present invention relates to a proximity-probe based detection assay for detecting an analyte in a sample and in particular to a method that comprises the use of at least one set of at least first and second proximity probes, which probes each comprise an analyte-binding domain and a nucleic acid domain and can simultaneously bind to the analyte directly or indirectly, wherein the nucleic acid domain of at least one of said proximity probes comprises a hairpin structure that can be unfolded by cleavage of the nucleic acid domain to generate at least one ligatable free end or region of complementarity to another nucleic acid molecule in said sample, wherein when the probes bind to said analyte unfolding said hairpin structure allows the nucleic acid domains of said at least first and second proximity probes to interact directly or indirectly.
摘要翻译： 本发明涉及一种用于检测样品中分析物的基于接近探针的检测测定法，特别涉及一种包括使用至少一组至少第一和第二接近探针的方法，该探针各自包含分析物 - 结合结构域和核酸结构域，并且可以直接或间接地同时结合分析物，其中至少一个所述邻近探针的核酸结构域包含发夹结构，所述发夹结构可以通过切割核酸结构域而被解折叠以产生至少 一个可连接的自由端或与所述样品中的另一个核酸分子互补的区域，其中当所述探针结合到所述分析物上展开所述发夹结构时，允许所述至少第一和第二接近探针的核酸结构域直接或间接相互作用。
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