公开(公告)号：US20190324033A1

公开(公告)日：2019-10-24

申请号：US15958931

申请日：2018-04-20

申请人： TOKITAE LLC ,  Cornell University

发明人： Luis F. Alonzo ,  Anne-Laure M. Le Ny ,  Kevin Paul Flood Nichols ,  Spencer Garing ,  Damian Madan ,  Sam Rasmussen Nugen ,  Troy Hinkley

IPC分类号： G01N33/569 ,  C12N15/73 ,  G01N33/543 ,  C12N7/02 ,  C12Q1/10 ,  C12Q1/66

摘要： Generally, this disclosure relates to expression constructs that encode a reporter enzyme-affinity binding tag fusion protein that is produced after the construct is inserted into bacteriophage and the bacteriophage infects bacteria. In some embodiments, the fusion protein is captured and produces a detectable signal. Signal intensity may correlate with the number of bacterial cells in a fluid sample. Methods of detecting bacteria using the expression constructs, and microfluidic devices for detecting bacteria using the expression constructs are also disclosed.

公开(公告)号：US20190217293A1

公开(公告)日：2019-07-18

申请号：US15870370

申请日：2018-01-12

申请人： Tokitae LLC

发明人： Luis F. Alonzo ,  Spencer Garing ,  Anne-Laure M. Le Ny ,  Kevin Paul Flood Nichols ,  Sam Rasmussen Nugen ,  John R. Williford

IPC分类号： B01L3/00 ,  B01D39/18 ,  G01N1/40 ,  C12Q1/04

CPC分类号： B01L3/502753 ,  B01D39/18 ,  B01D2239/1216 ,  B01L3/502738 ,  B01L2300/0681 ,  C12Q1/04 ,  G01N1/4077 ,  G01N2001/4088

摘要： A microfluidic device for concentrating and detecting bacteria in liquids, and related methods are described. The device includes a first filter chamber for capturing bacteria and performing incubations of the bacteria with one or more reagents, and a second filter chamber for capturing and concentrating a detectable material, with little or no binding of detectable material by the first filter. In an aspect, bacteria are incubated with growth media and engineered phage that cause the bacteria to produce an enzyme. In an aspect, the enzyme is capture in the second filter chamber and exposed to a substrate to produce a detectable signal.

公开(公告)号：US11608519B2

公开(公告)日：2023-03-21

申请号：US16049460

申请日：2018-07-30

申请人： TOKITAE LLC

发明人： Ted A. Baughman ,  Damian Madan ,  Eric Nalefski ,  Anne-Laure M. Le Ny

IPC分类号： C12Q1/6832 ,  C12N9/22 ,  C12N15/11

摘要： Embodiments disclosed herein include devices, methods, and systems for direct, selective, and sensitive detection of single-stranded and double-stranded target DNA sequences from various sources using a Cas12a protein. When activated by binding a target DNA sequence, the Cas12a cleaves a tether releasing a reporter molecule that may then be detected. In some embodiments, the systems, methods, and devices may include a filter or membrane that may help to separate the tethered and untethered reporter molecules. These devices, systems, and techniques allow a user to rapidly process samples that may contain the target DNA, without needing to amplify the target sequences. These devices and methods may be used to assay a wide variety of samples and target DNA sources, for the presence or absence of target DNA sequences. Compositions and kits, useful in practicing these methods, for example detecting a target DNA in a biological sample, are also described.

公开(公告)号：US20220106647A1

公开(公告)日：2022-04-07

申请号：US17430235

申请日：2020-02-07

申请人： Tokitae LLC

发明人： Ted Baughman ,  Anne-Laure M. Le Ny ,  Philip Leung ,  Damian Madan ,  Eric Nalefski

IPC分类号： C12Q1/6897 ,  C12N9/22 ,  C12N15/11

摘要： Embodiments disclosed herein include devices, methods, and systems for direct, selective, and sensitive detection of single-stranded and double-stranded target nucleic acid sequences from various sources in a solution-based system. When activated by binding a target nucleic acid sequence, the Cas protein cleaves a tether separating a reporter molecule from a capture moiety. The capture moiety can then be used to remove, localize, or sequester uncleaved molecule containing intact tethers. In some embodiments, the systems, methods, and devices may include a filter, a membrane, or other molecules that may help to separate the tethered and untethered reporter molecules and/or capture the reporter molecules. These devices, systems, and techniques allow a user to rapidly process samples that may contain the target nucleic acid, in some cases, without needing to amplify the target sequences, and without the need for sophisticated or expensive laboratory equipment. These devices and methods may be used to assay a wide variety of samples and target nucleic acid sources, for the presence or absence of a specific target sequences. Compositions and kits, useful in practicing these methods, for example detecting a target RNA or DNA in a biological sample, are also described.

公开(公告)号：US20200032325A1

公开(公告)日：2020-01-30

申请号：US16049460

申请日：2018-07-30

申请人： TOKITAE LLC

发明人： Ted A. Baughman ,  Damian Madan ,  Eric Nalefski ,  Anne-Laure M. Le Ny

IPC分类号： C12Q1/6832 ,  C12N9/22 ,  C12N15/11

摘要： Embodiments disclosed herein include devices, methods, and systems for direct, selective, and sensitive detection of single-stranded and double-stranded target DNA sequences from various sources using a programmed Cas12a protein. When activated by binding a target DNA sequence, the Cas12a cleaves a tether releasing a reporter molecule that may then be detected. In some embodiments, the systems, methods, and devices may include a filter or membrane that may help to separate the tethered and untethered reporter molecules. These devices, systems, and techniques allow a user to rapidly process samples that may contain the target DNA, without needing to amplify the target sequences. These devices and methods may be used to assay a wide variety of samples and target DNA sources, for the presence or absence of a specific target DNA sequences. Compositions and kits, useful in practicing these methods, for example detecting a target DNA in a biological sample, are also described.

公开(公告)号：US20200032324A1

公开(公告)日：2020-01-30

申请号：US16049457

申请日：2018-07-30

申请人： TOKITAE LLC ,  THE REGENTS OF THE UNIVERSITY OF CALIFORNIA

发明人： Ted A. Baughman ,  James Cate ,  Jennifer Doudna ,  Gavin John Knott ,  Damian Madan ,  Eric Nalefski ,  Anne-Laure M. Le Ny ,  Brittney Thornton

IPC分类号： C12Q1/6832 ,  C12N9/22 ,  C12N15/11

摘要： Embodiments disclosed herein include devices, methods, and systems for direct, selective, and sensitive detection of single-stranded target RNA sequences from various sources using a programmed Cas13a protein. When activated by binding a target RNA sequence, the Cas13a cleaves a tether releasing a reporter molecule that may then be detected. In some embodiments, the systems, methods, and devices may include a filter or membrane that may help to separate the tethered and untethered reporter molecules. These devices, systems, and techniques allow a user to rapidly process samples that may contain the target RNA, without needing to amplify the target sequences. These devices and methods may be used to assay a wide variety of samples and target RNA sources, for the presence or absence of a specific target RNA sequence. Compositions and kits, useful in practicing these methods, for example detecting a target RNA in a biological sample, are also described.