公开(公告)号：US5712476A

公开(公告)日：1998-01-27

申请号：US624789

申请日：1996-03-27

申请人： John A. Renfrew ,  James W. Cassidy

发明人： John A. Renfrew ,  James W. Cassidy

IPC分类号： G01N27/447 ,  G01N30/86 ,  H01J40/14

CPC分类号： G01N27/44721 ,  G01N30/8603

摘要： An improved electrophoresis and fluorescence detection apparatus has an electromagnetic radiation sensor juxtaposed with a sensing region. The output signal from the electromagnetic radiation sensor is a current signal, and the current signal is converted to a voltage signal. The voltage signal is summed with a programmable offset generated by an inexpensive eight-bit D/A converter. The offset signal is selected to establish a lower starting point for the dynamic range of the A/D conversion, and is selected to null some or all of the background electromagnetic radiation level. The summed signal is amplified and integrated in an integrator. The integrator is switchable under program control. The integrator is switched on for long and short intervals. The short intervals permit sensing over a dynamic range accommodating very high levels of fluorescence; very high peaks may be measured and features of the peaks distinguished. The long intervals permit sensing over a dynamic range that is optimized for the peaks associated with the smaller peaks of individual nucleotides. In this way, the dynamic range of the A/D convertor is set to permit the highest possible resolution over the range of interest during the time in which the sequencing of the nucleotides takes place. The sequencer is fast and economical and yields data with high resolution.

摘要翻译： 改进的电泳和荧光检测装置具有与感测区域并置的电磁辐射传感器。 来自电磁辐射传感器的输出信号是电流信号，电流信号被转换为电压信号。 电压信号与由便宜的8位D / A转换器产生的可编程偏移相加。 选择偏移信号以建立A / D转换的动态范围的较低起始点，并且被选择为零部分或全部背景电磁辐射水平。 求和信号被放大并集成在积分器中。 集成商可在程序控制下切换。 积分器长时间和短的间隔打开。 短时间间隔允许在适应非常高水平荧光的动态范围内进行感测; 可以测量非常高的峰，并区分峰的特征。 长间隔允许感测针对与各个核苷酸的较小峰相关的峰优化的动态范围。 以这种方式，A / D转换器的动态范围被设置为允许在发生核苷酸的测序的时间期间在感兴趣的范围内具有最高可能的分辨率。 音序器是快速和经济的，并产生高分辨率的数据。

公开(公告)号：US5786142A

公开(公告)日：1998-07-28

申请号：US452719

申请日：1995-05-30

申请人： John A. Renfrew ,  James W. Cassidy

发明人： John A. Renfrew ,  James W. Cassidy

IPC分类号： G01N27/447 ,  G01N30/86 ,  C12Q1/68 ,  B01D61/42 ,  C25B1/00 ,  G01N15/06

CPC分类号： G01N27/44721 ,  G01N30/8603

摘要： An improved electrophoresis and fluorescence detection method for nucleotide sequences comprises a fluorescence sensing region along the path of nucleotide detection, coupled with amplification and integration in an integrator of output signals in the form of activity peaks. The output signal, which is converted to a voltage signal, is summed with a programmable offset generated by an inexpensive eight-bit D/A converter. The offset signal is selected to establish a lower starting point for the dynamic range of analog-digital conversion, and is selected to null some or all of the background fluorescence level. The integrator is switchable under program control. The integrator is switched on for long and short integration intervals. The short intervals permit sensing over a dynamic range accommodating very high levels of fluorescence; very high peaks may be measured and features of the peaks distinguished. The long intervals permit sensing over a dynamic range that is optimized for the peaks associated with the smaller peaks of individual nucleotides. In this way, the dynamic range of the analog-digital conversion permits the highest possible resolution over the range of interest during the time in which the sequencing of the nucleotides takes place. The method of nucleotide sequencing and analysis is fast, economical, and yields data with high resolution.

摘要翻译： 用于核苷酸序列的改进的电泳和荧光检测方法包括沿着核苷酸检测途径的荧光感测区域，以及在活性峰形式的输出信号的积分器中的放大和积分。 转换为电压信号的输出信号与由廉价的八位D / A转换器产生的可编程偏移相加。 选择偏移信号以建立模拟数字转换的动态范围的较低起始点，并选择为零部分或全部背景荧光水平。 集成商可在程序控制下切换。 积分器接通长和短的积分间隔。 短时间间隔允许在适应非常高水平荧光的动态范围内进行感测; 可以测量非常高的峰，并区分峰的特征。 长间隔允许感测针对与各个核苷酸的较小峰相关联的峰优化的动态范围。 以这种方式，模拟数字转换的动态范围允许在发生核苷酸测序的时间内在感兴趣的范围内达到最高可能的分辨率。 核苷酸测序和分析的方法快速，经济，并且产生高分辨率的数据。

公开(公告)号：US6014213A

公开(公告)日：2000-01-11

申请号：US819910

申请日：1997-03-18

申请人： Paul Waterhouse ,  Alexandre M. Izmailov ,  Henryk Zaleski ,  John A. Renfrew ,  James W. Cassidy

发明人： Paul Waterhouse ,  Alexandre M. Izmailov ,  Henryk Zaleski ,  John A. Renfrew ,  James W. Cassidy

IPC分类号： G01N27/447 ,  G01N30/86 ,  G01N21/00

CPC分类号： G01N30/8603 ,  G01N27/44721

摘要： A high dynamic range apparatus for separation and detection of polynucleotide fragments has a housing adapted to receive an electrophoresis gel holder containing an electrophoresis gel loaded with fluorophore-labeled samples; one or more laser diodes for providing radiation of a frequency suitable for excitation of the fluorophore which irradiates a an array of excitation/detection sites on the electrophoresis gel; an array of detectors aligned with the excitation/detection sites for collecting fluorescent emissions; and one or more components for increasing the dynamic range of the instrument by at least an order of magnitude. These components, which can be used individually or in combination include detectors that are connected to a signal processing system that modulates the period of signal integration employed so that large signals are totaled at short time intervals and smaller signals are totaled at longer time intervals; the use of a beam splitter to produces a high intensity beam of emitted light and a low intensity beam of emitted light from each excitation/detection site; and a device for modulating the intensity of the excitation beam can be used to improve the dynamic range of the instrument.

摘要翻译： 用于分离和检测多核苷酸片段的高动态范围装置具有适于接收含有装载有荧光团标记的样品的电泳凝胶的电泳凝胶保持器的壳体; 用于提供适于激发荧光团的频率的辐射的一个或多个激光二极管，其照射电泳凝胶上的激发/检测位点阵列; 与用于收集荧光发射的激发/检测部位对准的检测器阵列; 以及用于将仪器的动态范围增加至少一个数量级的一个或多个组件。 可以单独或组合使用的这些组件包括连接到信号处理系统的检测器，该信号处理系统调制所采用的信号积分的周期，以便以较短的时间间隔合计大信号，并以更长的时间间隔合计较小的信号; 使用分束器产生高强度发射光束和来自每个激发/检测部位的低强度发射光束; 并且可以使用用于调制激发光束的强度的装置来改善仪器的动态范围。

公开(公告)号：US6136172A

公开(公告)日：2000-10-24

申请号：US470024

申请日：1999-12-22

申请人： John A. Renfrew ,  Eric Steinbach ,  Stuart MacMillan

发明人： John A. Renfrew ,  Eric Steinbach ,  Stuart MacMillan

IPC分类号： G01N27/447 ,  G01N27/26

CPC分类号： G01N27/44704 ,  G01N27/44743

摘要： Electrophoresis gels are formed using a gel-forming insert having a beveled edge which results in loading sites having a beveled bottom surface. The gel forming insert can have a continuous beveled edge across the entire width of the gel, in which case a special loading insert is used which matches the bevel of the gel. Alternatively, the gel forming insert may be formed with a plurality of fingers with beveled ends, each finger defining a well in the gel. In one form of the gel-forming insert, the fingers are formed from a soft, flexible polymer such as silicone applied on a rigid support.

公开(公告)号：US5710628A

公开(公告)日：1998-01-20

申请号：US353932

申请日：1994-12-12

申请人： Paul Waterhouse ,  John A. Renfrew ,  John K. Stevens

发明人： Paul Waterhouse ,  John A. Renfrew ,  John K. Stevens

IPC分类号： G01N21/64 ,  G01N27/447 ,  G01N21/00

CPC分类号： G01N27/44721

摘要： Improved detection methods and apparatus which may be used individually or in various combinations enhance the ability of the electrophoresis apparatus to detect fluorophore-labeled materials in short periods of time. One such apparatus comprises a housing adapted to receive an electrophoresis gel holder; an excitation source of electromagnetic radiation having a frequency effective to induce emission of electromagnetic radiation from the fluorophore; a plurality of optical fibers for delivering electromagnetic radiation from the excitation source to a linear array of excitation/detection sites on the gel holder, optical switching means for sequentially directing electromagnetic radiation into one of several pre-defined groups of the optical fibers; detection means such as a photomultiplier tube, or an array of photomultiplier tubes for detecting emission from the fluorophore induced by a radiation from the excitation source; and means for correlating a detected emission with the switching of the excitation electromagnetic radiation such that a given emission may be linked with the excitation/detection site being irradiated.

摘要翻译： 可以单独或以各种组合使用的改进的检测方法和装置增强电泳装置在短时间内检测荧光团标记的材料的能力。 一种这样的设备包括适于接收电泳凝胶保持器的壳体; 电磁辐射的激发源具有有效地诱导来自荧光团的电磁辐射的发射的频率; 用于将来自激发源的电磁辐射传递到凝胶保持器上的激发/检测部位的线性阵列的多个光纤，用于将电磁辐射顺序地引导到几个预定义的光纤组中的一个中的光开关装置; 检测装置，例如光电倍增管或用于检测由来自激发源的辐射引起的荧光团发射的光电倍增管阵列; 以及用于将检测到的发射与激发电磁辐射的切换相关联的装置，使得给定的发射可以与被照射的激发/检测部位相关联。

公开(公告)号：US5627022A

公开(公告)日：1997-05-06

申请号：US332577

申请日：1994-11-01

申请人： John A. Renfrew ,  Paul Waterhouse ,  John K. Stevens

发明人： John A. Renfrew ,  Paul Waterhouse ,  John K. Stevens

IPC分类号： G01N27/447 ,  C12Q1/00 ,  C12M1/40

CPC分类号： G01N27/44743 ,  G01N27/44704 ,  G01N27/44747

摘要： An electrophoresis microgel is formed in a gel holder. The gel holder comprises a top substrate, a bottom substrate and a spacer disposed between the top substrate and the bottom substrate. The spacer establishes a separation of from 25 to 250 microns between the top substrate and the bottom substrate. A gel compartment is formed by partially sealing the top substrate to the bottom substrate, while leaving an opening for the introduction of unpolymerized gel. The gel compartment is then filled with an unpolymerized gel, which is polymerized in the gel compartment. Electrodes may be printed on the substrates, may be contacts to an exposed edge of gel, or may be applied through windows cut into one of the substrates. One type of gel holder makes use of graded beads having a diameter of 25 to 250 microns slurried in an adhesive such as an acrylate adhesive as the spacer. The slurry is printed onto the surface of one or both substrates to form a spacer of the desired shape, and then hardened using heat or light. If desired, the spacer can establish lanes within the gel.

摘要翻译： 在凝胶保持器中形成电泳微凝胶。 凝胶保持器包括顶部基底，底部基底和设置在顶部基底和底部基底之间的间隔物。 间隔物在顶部基底和底部基底之间建立了从25至250微米的间隔。 通过将顶部基底部分地密封到底部基底而形成凝胶室，同时留下用于引入未聚合凝胶的开口。 然后用凝胶隔室填充未聚合的凝胶，其在凝胶隔室中聚合。 电极可以印刷在基底上，可以与凝胶的暴露边缘接触，或者可以通过切割成一个基底的窗口来施加。 一种类型的凝胶保持器使用在诸如丙烯酸酯粘合剂的粘合剂中作为间隔物浆化的直径为25至250微米的分级珠粒。 将浆料印刷到一个或两个基材的表面上以形成所需形状的间隔物，然后使用热或光硬化。 如果需要，间隔物可以在凝胶内建立泳道。

公开(公告)号：US5507934A

公开(公告)日：1996-04-16

申请号：US332892

申请日：1994-11-01

申请人： John A. Renfrew

发明人： John A. Renfrew

IPC分类号： G01N27/447 ,  G01N27/26

CPC分类号： G01N27/44704

摘要： An apparatus for the rapid filling and polymerization of electrophoresis gels comprises:(a) a housing;(b) a filling fixture removably disposed within the housing and adapted to receive a gel holder having an internal gel compartment;(c) an injection system, which is connectible to a reservoir for holding a polymerizable solution;(d) a solution injection connector adapted to couple the injection system to a gel holder placed within the filling fixture,(e) a controller for the injection system, which causes the injection system to inject polymerizable solution from the reservoir into the gel compartment; and(f) a radiation source disposed within the housing in a location effective to irradiate polymerizable solution within the gel compartment of a gel holder in the filling fixture. The filling fixture may be mounted on a base which is slidable between a position inside the housing, and a position outside the housing to permit easy placement of a gel holder into the filling fixture.

摘要翻译： 用于电泳凝胶的快速填充和聚合的装置包括：（a）壳体; （b）可移除地设置在所述壳体内并适于容纳具有内部凝胶隔室的凝胶保持器的填充固定器; （c）可连接到用于保持可聚合溶液的储存器的注射系统; （d）溶液注入连接器，其适于将注射系统耦合到置于填充夹具内的凝胶保持器，（e）用于注射系统的控制器，其使得注射系统将可聚合溶液从储存器注入凝胶室; 和（f）辐射源，其设置在壳体内的有效照射填充夹具中的凝胶保持器的凝胶室内的可聚合溶液的位置。 填充夹具可以安装在基座上，该基座可以在壳体内部的位置和壳体外部的位置之间滑动，以允许将凝胶保持器容易地放置到填充夹具中。

公开(公告)号：US6110344A

公开(公告)日：2000-08-29

申请号：US885531

申请日：1997-06-30

申请人： John A. Renfrew ,  Eric Steinbach ,  Stuart MacMillan

发明人： John A. Renfrew ,  Eric Steinbach ,  Stuart MacMillan

IPC分类号： G01N27/447 ,  G01N27/26

CPC分类号： G01N27/44704

摘要： Electrophoresis gels are formed using a gel-forming insert having a beveled edge which results in loading sites having a beveled bottom surface. The gel forming insert can have a continuous beveled edge across the entire width of the gel, in which case a special loading insert is used which matches the bevel of the gel. Alternatively, the gel forming insert may be formed with a plurality of fingers with beveled ends, each finger defining a well in the gel. In one form of the gel-forming insert, the fingers are formed from a soft, flexible polymer such as silicone applied on a rigid support.

摘要翻译： 使用具有倾斜边缘的凝胶形成插入物形成电泳凝胶，其导致具有斜面底表面的装载位置。 凝胶形成插入件可以在凝胶的整个宽度上具有连续的斜边，在这种情况下，使用与凝胶的斜面相匹配的特殊加载插入物。 或者，凝胶形成插入物可以形成有多个具有倾斜端部的指状物，每个手指在凝胶中限定孔。 在凝胶形成插入物的一种形式中，指状物由柔软的柔性聚合物（例如施加在刚性支撑体上的硅树脂）形成。

公开(公告)号：US6007689A

公开(公告)日：1999-12-28

申请号：US836268

申请日：1997-05-01

申请人： John A. Renfrew ,  Eric Steinbach ,  John K. Stevens ,  Henryk Zaleski

发明人： John A. Renfrew ,  Eric Steinbach ,  John K. Stevens ,  Henryk Zaleski

IPC分类号： G01N27/447 ,  G01N27/26

CPC分类号： G01N27/44704

摘要： An apparatus for the rapid preparation of electrophoresis gels comprises: (a) a housing; (b) a support fixture removably disposed within the housing and adapted to receive a gel holder having an internal gel compartment, the support fixture being optionally adapted to permit filling of the gel holder within the housing; (c) an optional injection system, which is connectible to a reservoir for holding a polymerizable solution; (d) an optional solution injection connector adapted to couple the injection system to a gel holder placed within the filling fixture, (e) an optional controller for the injection system, which causes the injection system to inject polymerizable solution from the reservoir into the gel compartment; and (f) a radiation source disposed within the housing in a location effective to irradiate polymierizable solution within the gel compartment of a gel holder in the support fixture. The support fixture may be mounted on a drawer which is slidable between a position inside the housing, and a position outside the housing to permit easy placement of a gel holder into the support fixture.

摘要翻译： PCT No.PCT / US95 / 13955 Sec。 371日期1997年5月1日 102（e）日期1997年5月1日PCT提交1995年10月31日PCT公布。 出版物WO96 / 日期：1996年5月9日。一种用于快速制备电泳凝胶的设备包括：（a）壳体; （b）可移除地设置在所述壳体内并适于接收具有内部凝胶隔室的凝胶保持器的支撑固定件，所述支撑固定件可选地适于允许在所述壳体内填充所述凝胶保持器; （c）可选的注射系统，其可连接到储存器以保持可聚合溶液; （d）可选的溶液注入连接器，其适于将注射系统耦合到置于填充夹具内的凝胶保持器，（e）用于注射系统的可选控制器，其使注射系统将可聚合溶液从储存器注入凝胶 隔间 以及（f）辐射源，其设置在所述壳体内的有效照射所述支撑夹具中的凝胶保持器的凝胶室内的可聚合溶液的位置。 支撑固定件可以安装在可在壳体内的位置和壳体外部的位置之间滑动的抽屉上，以允许将凝胶保持器容易地放置到支撑夹具中。

公开(公告)号：US6005663A

公开(公告)日：1999-12-21

申请号：US849662

申请日：1997-08-18

申请人： Paul Waterhouse ,  John A. Renfrew ,  John K. Stevens

发明人： Paul Waterhouse ,  John A. Renfrew ,  John K. Stevens

IPC分类号： G01N21/64 ,  G01N27/447 ,  G01N21/00

CPC分类号： G01N27/44721

摘要： Improved detection methods and apparatus which may be used individually or in combinations enhance the ability of the electrophoresis apparatus to detect fluorophore-labeled materials in short periods of time. One such apparatus comprises a housing adapted to receive an electrophoresis gel holder; an excitation source of electromagnetic radiation having a frequency effective to induce emission of electromagnetic radiation from the fluorophore; a plurality of optical fibers for delivering electromagnetic radiation from the excitation source to a linear array of excitation/detection sites on the gel holder, optical switching means for sequentially directing electromagnetic radiation into one of several pre-defined groups of the optical fibers; detection means such as a photomultiplier tube, or an array of photomultiplier tubes for detecting emission from the fluorophore induced by a radiation from the excitation source; and means for correlating a detected emission with the switching of the excitation electromagnetic radiation such that a given emission may be linked with the excitation/detection site being irradiated. For example, the optical switching means may alternate between directing radiation from the source into every other optical fiber, or may provide radiation in rotation to every third or fourth fiber. Alternatively, a spot array generation grating can be used for dividing an incident beam of coherent radiation into an array of excitation beamlets and directing each excitation beamlets to an excitation/detection site on the electrophoresis gel. Light emitting diode disposed to deliver excitation energy to an array of excitation/detection sites may also be used. This latter form of the apparatus is particularly advantageous due to the low costs of light emitting diodes (LEDs) compared to coherent light sources (e.g. lasers).