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1. 发明申请

US20110052037A1 Methods and Algorithms For Cell Enumeration in a Low-Cost Cytometer 有权
标题翻译：低成本细胞计数器细胞计数的方法和算法
公开(公告)号：US20110052037A1
公开(公告)日：2011-03-03
申请号：US12707578
申请日：2010-02-17
申请人： Frank A.W. Coumans , Jan Greve , Frank P. Modica , Leon W.M.M. Terstappen , Arjan G.J. Tibbe , John A. Verrant
发明人： Frank A.W. Coumans , Jan Greve , Frank P. Modica , Leon W.M.M. Terstappen , Arjan G.J. Tibbe , John A. Verrant
IPC分类号： G06K9/00
CPC分类号： G01N15/1475 , G01N33/5094 , G01N33/54333
摘要： The enumeration of cells in fluids by flow cytometry is widely used across many disciplines such as assessment of leukocyte subsets in different bodily fluids or of bacterial contamination in environmental samples, food products and bodily fluids. For many applications the cost, size and complexity of the instruments prevents wider use, for example, CD4 analysis in HIV monitoring in resource-poor countries. The novel device, methods and algorithms disclosed herein largely overcome these limitations. Briefly, all cells in a biological sample are fluorescently labeled, but only the target cells are also magnetically labeled. In addition, non-magnetically labeled cells are imaged for viability in a modified slide configuration. The labeled sample, in a chamber or cuvet, is placed between two wedge-shaped magnets to selectively move the magnetically labeled cells to the observation surface of the cuvet. An LED illuminates the cells and a CCD camera captures the images of the fluorescent light emitted by the target cells. Image analysis performed with a novel algorithm provides a count of the cells on the surface that can be related to the target cell concentration of the original sample. The compact cytometer system provides a rugged, affordable and easy-to-use technique, which can be used in remote locations.
摘要翻译：通过流式细胞术对流体细胞的计数被广泛应用于许多学科，例如评估不同体液中的白细胞亚群或环境样品，食品和体液中的细菌污染。对于许多应用，仪器的成本，尺寸和复杂性阻止了更广泛的使用，例如资源贫乏国家的艾滋病毒监测中的CD4分析。本文公开的新颖的装置，方法和算法在很大程度上克服了这些限制。简言之，生物样品中的所有细胞都被荧光标记，但只有靶细胞也被磁性标记。另外，非磁性标记的细胞在修改的载玻片配置中的存活力被成像。将标记的样品置于室或铜管中，放置在两个楔形磁体之间，以选择性地将磁性标记的细胞移动到铜管的观察表面。 LED照亮细胞，CCD相机捕获由靶细胞发射的荧光的图像。用新算法进行的图像分析提供了与原始样品的靶细胞浓度相关的表面上的细胞计数。紧凑型细胞仪系统提供坚固，经济实惠且易于使用的技术，可用于偏远地区。

2. 发明申请

US20110014686A1 Method and apparatus for imaging target components in a biological sample using permanent magnets 审中-公开
标题翻译：用于使用永磁体对生物样品中的目标成分进行成像的方法和装置
公开(公告)号：US20110014686A1
公开(公告)日：2011-01-20
申请号：US12888562
申请日：2010-09-23
申请人： Arjan G.J. Tibbe , Leon W.M.M. Terstappen
发明人： Arjan G.J. Tibbe , Leon W.M.M. Terstappen
IPC分类号： C12M1/42
CPC分类号： G01N33/582 , G01N33/54326 , G01N33/56972 , G01N2333/70514 , G01N2446/00
摘要： A system for enumeration of cells in fluids by image cytometry is described for assessment of target populations such as leukocyte subsets in different bodily fluids or bacterial contamination in environmental samples, food products and bodily fluids. Briefly, fluorescently labeled target cells are linked to magnetic particles or beads. In one embodiment, a small, permanent magnet is inserted directly into the chamber containing the labeled cells. The magnets are coated with PDMS silicone rubber to provide a smooth and even surface which allows imaging on a single focal plane. The magnet is removed from the sample and illuminated with fluorescent light emitted by the target cells captured by a CCD camera. In another embodiment, a floater having a permanent magnet allows the target cells to line up along a single imaging plane within the sample solution. Image analysis can be performed with a novel algorithm to provide a count of the cells on the surface, reflecting the target cell concentration of the original sample.
摘要翻译：描述了通过图像细胞计数法检测流体细胞的系统，用于评估目标群体，例如不同体液中的白细胞亚群或环境样品，食品和体液中的细菌污染物。简言之，荧光标记的靶细胞与磁性颗粒或珠粒连接。在一个实施例中，将小的永磁体直接插入到包含标记电池的腔室中。这些磁体用PDMS硅橡胶涂覆，以提供平滑和均匀的表面，允许在单个焦平面上进行成像。从样品中除去磁体，并用由CCD照相机俘获的靶细胞发射的荧光照射。在另一个实施例中，具有永磁体的浮子允许靶细胞沿着样品溶液内的单个成像平面排列。可以用新颖的算法进行图像分析，以提供表面上的细胞计数，反映原始样品的靶细胞浓度。

3. 发明申请

US20090061477A1 Method and apparatus for imaging target components in a biological sample using permanent magnets 有权
标题翻译：用于使用永磁体对生物样品中的目标成分进行成像的方法和装置
公开(公告)号：US20090061477A1
公开(公告)日：2009-03-05
申请号：US12031807
申请日：2008-02-15
申请人： Arjan G.J. Tibbe , Leon W.M.M. Terstappen
发明人： Arjan G.J. Tibbe , Leon W.M.M. Terstappen
IPC分类号： C12Q1/06 , C12M1/00
CPC分类号： G01N33/582 , G01N33/54326 , G01N33/56972 , G01N2333/70514 , G01N2446/00
摘要： A system for enumeration of cells in fluids by image cytometry is described for assessment of target populations such as leukocyte subsets in different bodily fluids or bacterial contamination in environmental samples, food products and bodily fluids. Briefly, fluorescently labeled target cells are linked to magnetic particles or beads. In one embodiment, a small, permanent magnet is inserted directly into the chamber containing the labeled cells. The magnets are coated with PDMS silicone rubber to provide a smooth and even surface which allows imaging on a single focal plane. The magnet is removed from the sample and illuminated with fluorescent light emitted by the target cells captured by a CCD camera. In another embodiment, a floater having a permanent magnet allows the target cells to line up along a single imaging plane within the sample solution. Image analysis can be performed with a novel algorithm to provide a count of the cells on the surface, reflecting the target cell concentration of the original sample.
摘要翻译：描述了通过图像细胞计数法检测流体细胞的系统，用于评估目标群体，例如不同体液中的白细胞亚群或环境样品，食品和体液中的细菌污染物。简言之，荧光标记的靶细胞与磁性颗粒或珠粒连接。在一个实施例中，将小的永磁体直接插入到包含标记电池的腔室中。这些磁体用PDMS硅橡胶涂覆，以提供平滑和均匀的表面，允许在单个焦平面上进行成像。从样品中除去磁体，并用由CCD照相机俘获的靶细胞发射的荧光照射。在另一个实施例中，具有永磁体的浮子允许靶细胞沿着样品溶液内的单个成像平面排列。可以用新颖的算法进行图像分析，以提供表面上的细胞计数，反映原始样品的靶细胞浓度。

4. 发明申请

US20120055854A1 Filter Method for Separating Unbound Ferrofluid from Target-bound Ferrofluid in a Biological Sample 审中-公开
标题翻译：将生物样品中未结合的铁磁流体与目标结合的铁磁流体分离的过滤方法
公开(公告)号：US20120055854A1
公开(公告)日：2012-03-08
申请号：US13147908
申请日：2010-02-05
申请人： Arjan G.J. Tibbe
发明人： Arjan G.J. Tibbe
IPC分类号： B03C1/02
CPC分类号： B03C1/286 , B03C1/01 , B03C2201/18
摘要： A filtering system for separating unbound ferrofluid from bound ferrofluid in the enrichment of a target entity in a biological sample. The filtering device of the present invention has application in the isolation target cells from unbound ferrofluid during separation with a permanent magnet (floater) mechanism. This process reduces interference of unbound ferrofluid during subsequent image analysis or enumeration of cells by image cytometry. The system has application in the assessment of target populations such as leukocyte subsets in different bodily fluids or bacterial contamination in environmental samples, food products and bodily fluids. Briefly, fluorescently labeled target cells are linked to magnetic particles or beads. The linkage process results in a mixture having a population of contaminating unbound magnetic particles. In one embodiment for separation, a small, permanent magnet is inserted directly into the chamber containing the labeled cells. The magnets are coated with PDMS silicone rubber to provide a smooth and even surface which allows imaging on a single focal plane. A filter is positioned on a cover of the floater device to allow unbound ferrofluid to pass through the pores, but restrict the passage of the target entity. The floater and filter are removed from the sample and the filter surface is illuminated with fluorescent light emitted by the target cells captured by a CCD camera. Image analysis can be performed with a novel algorithm to provide a count of the cells on the surface, reflecting the target cell concentration of the original sample.
摘要翻译：一种用于在生物样品中浓缩目标实体中分离未结合的铁磁流体与结合铁磁流体的过滤系统。本发明的过滤装置在用永久磁铁（浮子）机构分离的同时，可以将其从未结合的铁磁流体中分离出来。该过程减少未结合的铁磁流体在随后的图像分析或通过图像细胞计数细胞计数的干扰。该系统可用于评估目标人群，如不同体液中的白细胞亚群或环境样品，食品和体液中的细菌污染。简言之，荧光标记的靶细胞与磁性颗粒或珠粒连接。连接过程产生具有污染的未结合磁性颗粒群的混合物。在用于分离的一个实施例中，将小的永磁体直接插入到含有标记细胞的腔室中。这些磁体用PDMS硅橡胶涂覆，以提供平滑和均匀的表面，允许在单个焦平面上进行成像。过滤器定位在浮子装置的盖上，以允许未结合的铁磁流体穿过孔，但限制目标实体的通过。从样品中除去浮子和过滤器，并用由CCD照相机捕获的靶细胞发射的荧光照射滤光片表面。可以用新颖的算法进行图像分析，以提供表面上的细胞计数，反映原始样品的靶细胞浓度。

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