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1. 发明授权

US09982251B2 Large volume ex vivo electroporation method 有权
公开(公告)号：US09982251B2
公开(公告)日：2018-05-29
申请号：US10537254
申请日：2004-03-15
申请人： Richard E. Walters , Alan D. King
发明人： Richard E. Walters , Alan D. King
IPC分类号： C12N13/00 , C12N15/87 , C12M3/02 , C12M3/00 , C12M1/42 , C12M1/00 , A61K9/00
CPC分类号： C12N13/00 , A61K9/0009 , C12M1/42 , C12M3/00 , C12M3/006 , C12M3/02 , C12M35/02 , C12M45/07 , C12N15/87
摘要： An object of the invention is to provide an electroporation method for treating vesicles with exogenous material for insertion of the exogenous material into the vesicles which includes the steps of: a. retaining a suspension of the vesicles and the exogenous material in a treatment volume in a chamber which includes electrodes, wherein the chamber has a geometric factor (cm.sup.−1) defined by the quotient of the electrode gap squared (cm.sup.2) divided by the chamber volume (cm.sup.3), wherein the geometric factor is less than or equal to 0.1 cm.sup.−1, wherein the suspension of the vesicles and the exogenous material is in a medium which is adjusted such that the medium has conductivity in a range spanning 50 microSiemens/cm to 500 microSiemens/cm, wherein the suspension is enclosed in the chamber during treatment, and b. treating the suspension enclosed in the chamber with one or more pulsed electric fields. With the method, the treatment volume of the suspension is scalable, and the time of treatment of the vesicles in the chamber is substantially uniform.

2. 发明授权

US07824901B2 Non-uniform electric field chamber for cell fusion 有权
标题翻译：用于细胞融合的不均匀电场室
公开(公告)号：US07824901B2
公开(公告)日：2010-11-02
申请号：US10580429
申请日：2003-12-01
申请人： Richard E. Walters
发明人： Richard E. Walters
IPC分类号： C12M3/00 , C12M1/34
CPC分类号： C12N13/00 , C12M35/02
摘要： An apparatus is provided for carrying out fusion of biological cells (10) and includes a base member (24) on which a conductive outer electrode (18) is supported and has an outer electrode radius (r2) and has an electrode height (19). A conductive inner electrode (20) is supported on the base member (24) and has an inner electrode radius (r1) and also has the electrode height (19). The outer and inner electrodes (18,20) are spaced apart from each other by a gap which defines a fusion chamber (14). The inner electrode radius (r1), the outer electrode radius (r2), and the gap are selected in accordance with a predetermined range of selectable ratios (r1/r2) in a range from 0.7 to 0.9, wherein a selected gap is limited by the range of selectable ratios (r1/r2), and wherein a determined ratio (r1/r2) among the selectable ratios is based on the selected gap, such that compression between the biological cells (10) and permeability between cell membranes are maximized and temperature rise is minimized for providing cell fusion in the fusion chamber (14).
摘要翻译：提供一种用于进行生物电池（10）熔化的装置，并且包括一个支撑导电外电极（18）并具有外电极半径（r2）并且具有电极高度（19）的基体部件（24）。导电内电极（20）支撑在基体部件（24）上，内电极半径（r1）也具有电极高度（19）。外部和内部电极（18,20）通过限定熔化室（14）的间隙彼此间隔开。内部电极半径（r1），外部电极半径（r2）和间隙根据可选择比率（r1 / r2）的预定范围在0.7至0.9的范围内选择，其中所选择的间隙由可选择比率（r1 / r2）的范围，并且其中可选择比率之间的确定比率（r1 / r2）基于所选择的间隙，使得生物细胞（10）之间的压缩和细胞膜之间的渗透性最大化，使熔化室（14）中的细胞融合最小化温度上升。

3. 发明授权

US06603998B1 Delivery of macromolecules into cells 失效
标题翻译：将大分子递送到细胞中
公开(公告)号：US06603998B1
公开(公告)日：2003-08-05
申请号：US09889581
申请日：2001-07-30
申请人： Alan D. King , Richard E. Walters
发明人： Alan D. King , Richard E. Walters
IPC分类号： A61N130
CPC分类号： A61N1/325 , A61N1/306 , C12M35/02 , C12N15/87
摘要： An object of the invention is to provide a method for delivery of macromolecules into biological cells, such as Langerhans cells (22) in the epidermis (20) of a patient, which includes the steps of coating electodes (16) in an electrode assembly (12) with solid phase macromolecules to be delivered, such as a DNA, and/or RNA vaccine or a protein-based vaccine, attaching the electrode assembly (12) having the coated electrodes (16) to an electrode assembly holder (13), providing a waveform generator (15), establishing electrically conductive pathways between the electrodes (16), and the waveform generator (15), locating the electrodes (16) such that the biological cells are situated therebetween, such as by penetrating the needle electrode (16) into the epidermis (20) above the epidermal basal lamina, and providing pulse waveform from the waveform generator (15) to the electrodes (16), such that macromolecule on the electrodes (16) is driven off of the electrodes (16), and delivered into the biological cells, such as the Langerhans cells (22).
摘要翻译：本发明的目的是提供一种用于将大分子递送到生物细胞的方法，例如患者的表皮（20）中的朗格汉斯细胞（22），其包括在电极组件中涂覆电极（16）的步骤将具有涂覆电极（16）的电极组件（12）附接到电极组件保持器（13）上的要传递的固相大分子如DNA和/或RNA疫苗或基于蛋白质的疫苗，提供波形发生器（15），在电极（16）和波形发生器（15）之间建立导电通路，定位电极（16），使得生物细胞位于其间，例如穿过针电极（ 16）进入表皮基底层上方的表皮（20），并将波形发生器（15）的脉搏波形提供给电极（16），使得电极（16）上的大分子从电极（16）驱动，，并交付生物细胞，如朗格汉斯细胞（22）。

4. 发明授权

US6117660A Method and apparatus for treating materials with electrical fields having varying orientations 失效
标题翻译：用于处理具有不同取向的电场的材料的方法和装置
公开(公告)号：US6117660A
公开(公告)日：2000-09-12
申请号：US242174
申请日：1999-02-10
申请人： Richard E. Walters , Alan D. King , Derin C. Walters
发明人： Richard E. Walters , Alan D. King , Derin C. Walters
IPC分类号： C12N13/00
CPC分类号： C12N13/00 , C12M35/02
摘要： The object of the invention is to provide a method and apparatus for treating membrane containing material with electrical fields and with an added treating substance. With the method, a plurality of electrodes (121-128) are arrayed around the material to be treated and are connected to outputs of an electrode selection apparatus (110). Inputs of the electrode selection apparatus are connected to outputs of an agile pulse sequence generator. A treating substance is added to the membrane-containing material. Electrical pulses are applied to the electrode selection apparatus and are routed through the electrode selection apparatus in a predetermined, computer-controlled sequence to selected electrodes in the array of electrodes, whereby the membrane containing material is treated with the added treating substance and with electrical fields of sequentially varying directions. The routing of applied pulses through the electrode selection apparatus (110) to selected electrodes (121-128) can be done in an enormous number of ways.
摘要翻译： PCT No.PCT / US97 / 09300 Sec。 371日期1999年2月10日 102（e）1999年2月10日PCT 1997年6月10日PCT公布。第WO98 / 56893号公报日期：1998年12月17日本发明的目的是提供一种用电场和附加处理物质处理含膜材料的方法和装置。利用该方法，将多个电极（121-128）围绕待处理的材料排列并连接到电极选择装置（110）的输出端。电极选择装置的输入连接到敏捷脉冲序列发生器的输出端。将处理物质加入到含膜材料中。将电脉冲施加到电极选择装置，并以预定的计算机控制的顺序通过电极选择装置引导到电极阵列中的选定电极，由此将含有膜的材料用添加的处理物质和电场顺序变化的方向。施加的脉冲通过电极选择装置（110）到选定电极（121-128）的布线可以以许多方式进行。

5. 发明授权

US4147120A Auxiliary feed mechanism for sewing machines 失效
标题翻译：缝纫机辅助进给机构
公开(公告)号：US4147120A
公开(公告)日：1979-04-03
申请号：US833110
申请日：1977-09-14
申请人： Maximilian Adamski, Jr. , Robert C. Talsma , Richard E. Walters
发明人： Maximilian Adamski, Jr. , Robert C. Talsma , Richard E. Walters
IPC分类号： D05B27/12 , D05B21/00
CPC分类号： D05B27/12 , D05D2207/02
摘要： An auxiliary feed mechanism adapted for use with a sewing machine including a feed roller for advancing the workpiece through the work station and a mounting structure adapted to move the feed roller between two positions in one of which the feed roller is operative to aid in feeding the workpiece through the workstation and in the other of which the feed roller is rendered ineffective to transmit motive power to the workpiece. A plurality of flexible couplings drive the feed roller in timed relation to the actuation of a variable speed drive motor which is coupled in a unique electric circuit relationship with the main drive motor of the machine so as to vary the feeding rate of the feed roller in accordance with the machine cycle rate.
摘要翻译：一种适于与缝纫机一起使用的辅助进给机构，该缝纫机包括用于使工件通过工作台前进的进给辊和适于将进给辊移动到两个位置之间的安装结构，其中一个进给辊可操作以帮助进给工件通过工作站，另一个工件使进给辊无效地将动力传递到工件。多个柔性联轴器以与驱动机构的驱动电动机独特的电路关系的变速驱动电动机的动作的定时关系驱动进给辊，以便改变进给辊的进给速度按照机器循环速率。

6. 发明申请

US20110166501A1 Method and apparatus for the delivery of polynucleotide vaccines to mammalian skin 审中-公开
标题翻译：用于将哺乳动物皮肤递送多核苷酸疫苗的方法和装置
公开(公告)号：US20110166501A1
公开(公告)日：2011-07-07
申请号：US13065135
申请日：2011-03-15
申请人： Richard E. Walters , Derin C. Walters , Alan D. King , Anna-Karin Roos
发明人： Richard E. Walters , Derin C. Walters , Alan D. King , Anna-Karin Roos
IPC分类号： A61M37/00
CPC分类号： A61N1/327 , A61K39/12 , A61K2039/53 , A61K2039/54 , A61K2039/70 , A61N1/306 , C07K14/005 , C12N2770/24122 , C12N2770/24134
摘要： An object of the invention is to provide a method and apparatus for the delivery of polynucleotide vaccines into mammalian skin cells to increase T cell response and to reduce pain and discomfort due to long electric waveform application and due to muscle contractions. The method for the delivery of polynucleotide vaccines into mammalian skin cells includes the steps of: (a.) administering a polynucleotide vaccine into the skin at an administration site, (b.) applying a needle electrode to the skin in the vicinity to the administration site, and (c.) applying a sequence of at least three single, operator-controlled, independently programmed, narrow interval electrical waveforms, which have pulse intervals that are less than 100 milliseconds, to deliver the polynucleotide vaccine into the skin cells by electroporation. The sequence of at least three waveforms has one, two, or three of the following characteristics (1) at least two of the at least three waveforms differ from each other in waveform amplitude, (2) at least two of the at least three waveforms differ from each other in waveform width, and (3) a first waveform interval for a first set of two of the at least three waveforms is different from a second waveform interval for a second set of two of the at least three waveforms.
摘要翻译：本发明的目的是提供一种用于将多核苷酸疫苗递送到哺乳动物皮肤细胞中以增加T细胞应答并减少由于长电波形应用引起的疼痛和不适以及由于肌肉收缩引起的疼痛和不适的方法和装置。用于将多核苷酸疫苗递送到哺乳动物皮肤细胞中的方法包括以下步骤：（a）在给药部位将多核苷酸疫苗施用于皮肤，（b）将针电极施用于给药的附近的皮肤站点，以及（c）应用脉冲间隔小于100毫秒的至少三个单个，操作者控制的，独立编程的窄间隔电波形序列，以通过电穿孔将多核苷酸疫苗递送到皮肤细胞中。至少三个波形的序列具有以下特征（1）中的一个，两个或三个，所述至少三个波形中的至少两个波形在波形幅度上彼此不同，（2）至少三个波形中的至少两个波形在波形宽度上彼此不同，以及（3）至少三个波形中的两个的第一组的第一波形间隔与用于至少三个波形中的两个的第二组的第二波形间隔不同。

7. 发明授权

US07371561B2 In vitro, multiple electrode pair array and multiple treatment cell apparatus for use in electroporation 有权
标题翻译：体外，多电极对阵列和用于电穿孔的多处理细胞装置
公开(公告)号：US07371561B2
公开(公告)日：2008-05-13
申请号：US10694345
申请日：2003-10-28
申请人： Richard E. Walters , Paul K. Gustavson
发明人： Richard E. Walters , Paul K. Gustavson
IPC分类号： C12M1/42
CPC分类号： C12M35/02 , B01L3/5085 , B01L2300/0829 , B01L2400/0415 , C12M23/12
摘要： A multiple electrode pair array apparatus is provided for use with a multiple well plate which has multiple wells distributed in a two-dimensional matrix array which has R rows and C columns. The multiple electrode pair array apparatus includes a non-conductive base member. An array of pairs of electrodes are attached to the base member and project therefrom. The pairs of electrodes are distributed on the base member in a two-dimensional matrix array which has R rows and C columns to enable registration with the two-dimensional matrix array of wells. Each pair of electrodes includes a respective first electrode and a respective second electrode. An array of R row conductors are electrically connected to corresponding first electrodes. An array of C column conductors are oriented perpendicular to the R row conductors, are electrically insulated from the row conductors, and are electrically connected to corresponding second electrodes.
摘要翻译：提供多电极对阵列装置用于多孔板，多孔板具有分布在具有R行和C列的二维矩阵阵列中的多个阱。多电极对阵列装置包括非导电基底部件。一对电极被附接到基底构件并从其突出。电极对以具有R行和C列的二维矩阵阵列分布在基座上，以使得能够与孔的二维矩阵阵列配准。每对电极包括相应的第一电极和相应的第二电极。 R排导体的阵列电连接到对应的第一电极。 C列导体的阵列垂直于R行导体定向，与行导体电绝缘，并且电连接到对应的第二电极。

8. 发明授权

US06878538B1 Apparatus for receiving and aligning a multiple electrode pair array and a multiple treatment cell for use in electroporation 有权
标题翻译：用于接收和对准多个电极对阵列和用于电穿孔的多重处理电池的装置
公开(公告)号：US06878538B1
公开(公告)日：2005-04-12
申请号：US10694344
申请日：2003-10-28
申请人： Richard E. Walters , Benjamin R. Lane , Michael S. Abbott
发明人： Richard E. Walters , Benjamin R. Lane , Michael S. Abbott
IPC分类号： B01L9/00 , C12M1/42 , C12M3/00 , C12N20060101
CPC分类号： B01L9/523 , B01L2300/046 , B01L2300/0829 , C12M23/12 , C12M23/48 , C12M35/02 , Y10S435/809
摘要： An apparatus is provided for receiving and aligning a multiple electrode pair array and a multiple well plate used in electroporation. The apparatus includes a housing which includes a bottom housing portion and a pair of side housing portions which project upward from the bottom housing portion. A pair of board-reception structures are adjacent to the inner portions of the side housing portions. Multiple well plate engaging and disengaging means, supported by the housing, are provided for engaging a multiple well plate with a multiple electrode pair array and for disengaging a multiple well plate from a multiple electrode pair array. Preferably, the multiple well plate engaging and disengaging means include a multiple well plate lifting and lowering assembly. The multiple well plate lifting and lowering assembly can include lifting/lowering handles located outside the housing.
摘要翻译：提供了用于接收和对准多电极对阵列和用于电穿孔的多孔板的装置。该装置包括壳体，该壳体包括底部壳体部分和从底部壳体部分向上突出的一对侧部壳体部分。一对板接收结构邻近侧壳体部分的内部部分。提供由壳体支撑的多个孔板接合和分离装置，用于使多孔板与多个电极对阵列接合并且用于使多个孔板与多个电极对阵列分离。优选地，多孔板接合和分离装置包括多孔板升降组件。多孔板升降组件可以包括位于壳体外部的升降手柄。

9. 发明授权

US06713291B2 Electrodes coated with treating agent and uses thereof 有权
公开(公告)号：US06713291B2
公开(公告)日：2004-03-30
申请号：US09920861
申请日：2001-08-03
申请人： Alan D. King , Richard E. Walters
发明人： Alan D. King , Richard E. Walters
IPC分类号： C12N1300
CPC分类号： A61N1/325 , A61H39/002 , A61N1/306
摘要： An object of the invention is to provide a method for delivery of macromolecules into biological cells in the tissues of a patient and includes the steps of: (a) providing electrodes (16) in an electrode assembly (12), wherein the electrodes have fixed electrode surfaces (42) which are coated with at least one static layer of electrode releasable molecules (44) to be delivered; (b) providing a waveform generator (15) for generating electric fields; (c) establishing electrically conductive pathways between the electrodes (16) and the waveform generator (15); (d) locating the electrodes (16) such that the biological cells are situated therebetween, and (g) providing electric fields in the form of pulse waveforms from the waveform generator (15) to the electrodes (16), such that molecules in the at least one static layer of the electrode releasable molecules (44) on the electrodes (16) are delivered into the biological cells. The electrode releasable molecules (44) can be either electric field separable molecules and/or solvent separable material. Another object of the invention is to provide an apparatus for carrying out the method of the invention. The static-coated electrode assembly (12) can be provided in a sterile package (24), from which the electrode assembly (12) is removed prior to use. The statically-coated electrode assembly (12) can be in a form of a disposable assembly (12) which is removable and replaceable from an electrode assembly holder (13).

10. 发明授权

US06653114B2 Method and apparatus for treating materials with electrical fields having varying orientations 失效
公开(公告)号：US06653114B2
公开(公告)日：2003-11-25
申请号：US10265133
申请日：2002-10-07
申请人： Richard E. Walters , Alan D. King , Derin C. Walters
发明人： Richard E. Walters , Alan D. King , Derin C. Walters
IPC分类号： C12N1300
CPC分类号： C12N13/00 , C12M35/02
摘要： The object of the invention is to provide a method and apparatus for treating membrane-containing material with electrical fields, in vitro, and with an added treating substance. With the method, a plurality of electrodes are arrayed around the material to be treated and are connected to outputs of an electrode selection apparatus. Inputs of the electrode selection apparatus are connected to outputs of a pulse sequence generator. A treating substance is added to the membrane-containing material. Electrical pulses are applied to the electrode selection apparatus and are routed through the electrode selection apparatus in a predetermined, computer-controlled sequence to selected electrodes in the array of electrodes, whereby the membrane-containing material is treated with the added treating substance and with electrical fields. The routing of applied pulses through the electrode selection apparatus to selected electrodes can be done in an enormous number of ways.

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