CN100458441C - 用于测量血液凝固的装置及其方法 - Google Patents
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Abstract
一种用于测量流体，特别是血液在微通道中的凝固时间的装置和方法，其中凝固的开始是由位于微通道的任何一侧上的两个电极之间的电容或者阻抗的变化率或者数值的测量来确定的。提供一种包括上支撑件(101)和下支撑件(103)的一次性测试带(100)。微通道(102)形成在下支撑件(103)的上表面中。第二基层(101)层压在支撑件(103)的顶部上，从而封闭开放的微通道(102)。电极(104)形成在基层的相应的外表面上并且与通道共面。
Description
技术领域
本发明涉及一种测量生物试样的凝固因子的装置和方法。本发明特别涉及一种可插入到能够显示凝固化验结果的手持式或者便携式测量仪器中的一次性使用的试片。这样一种装置适用于家庭测试或者护理处。
背景技术
人体中在血管损伤后抑制血液流动的能力对于持续生存是极为重要的。实现这种情况的方法被称为止血法并且是通过能够形成血凝块或者血栓的血液凝固作用实现的。血凝块是由陷在不可溶解的血纤蛋白颗粒网中的血小板栓构成的。尽管凝块的形成是必要的，这样的凝块的存留对于人体是危险的。这样，为了在凝固作用已经达到其目的后使人体受到的伤害达到最小，包围凝块的健康细胞释放纤溶酶以消化血纤蛋白，从而分解凝块。但是，在世界范围内，血栓症是导致死亡的一个主要原因，这是由于通向生命器官和组织的血液流动被血凝块堵塞。血栓可能出现在循环系统中的任何位置，但是，特别是当在这种情况出现在下半身、心脏、肺或者脑中可能导致深静脉血栓形成、急性心肌梗塞、肺栓塞或者急性缺血性中风时会危及生命。
导致凝块形成的两种路径或者凝血级联被称为内在路径和外在路径。这两种路径是由不同的机理启动的但沿着一个共同的路径会聚。在没有组织损伤的情况下，响应于异常的管壁形成凝块是内在路径的成因，而响应于组织损伤形成凝块是外在路径的成因。凝血级联是非常复杂的并且包括多个被称为凝固因子的不同蛋白质。
患有心脏或者血管疾病的人和动过外科手术的患者具有可能导致危急生命的临床状态的凝块形成的危险。这样的人通常需要利用降低血液粘稠性的药物或者抗凝血剂进行治疗。但是，血流中的抗凝血剂的量必须保持在适合的水平；太少可能导致不希望的凝固而太多可能导致出血。因此，已经开发了常规凝固测试以评价血液或者血浆的凝固状况。
一种有效的凝固测量是所谓的凝血酶原时间(PT)测试。PT测试是于1935年首次提出的并且测量血液或者血浆的组织诱导因子凝固时间。这可提供一种外在凝固路径的评价并且对于因子I、II、V、VII和X是敏感的。该测试是通过将诸如促凝血酶原激酶和Ca2+的凝血剂加入到患者试样中并且测量凝块形成的时间。已经开发了诸如CoaguChek
凝固测量仪器的便携式凝固监测器，这样的监测器利用来自于刺破手指或者切割装置的非抗凝固毛细管全血测量凝血酶原时间。这样的监测器已经被证明是一种适合长期口头抗凝血治疗患者的有价值的工具。
但是，PT测试结果的常规表示方式不适于国际间的比较，这是由于这些数值取决于所用促凝血酶原激酶的性质。这导致采用国际标准比率或者INR作为表示凝血酶原时间的一种形式，其中：
INR＝(PT ratio)ISI，其中ISI是国际灵敏度指数；以及
PT ratio＝患者的PT/平均正常PT
ISI是根据利用特定的促凝血酶原激酶相对于促凝血酶原激酶的世界卫生组织(WHO)国际标准参考制品(人类组合67/40)获得的多个试样的PT值的校准线导出的。考虑特定方法和所用促凝血酶原激酶类型的ISI的特定值被分配给每一个PT系统，从而可将每一个PT比率可以转变成标准化比率。通过使用INR，患者应该能够保持令人满意的凝固水平，这不取决于所用的PT系统。
测量在血液或者血浆中凝固的另一种方法是激活部分促凝血酶原激酶时间测试(APTT)。该测试是对在激活内在路径时出现的凝固时间的测量。在存在钙离子和磷脂(部分促凝血酶原激酶)时，可通过将激活剂(高岭土)添加到试样中来实现。利用APTT评价包括因子I、II、V、VIII、X、XI和XII的内在凝固路径。在磷脂表面上形成复合体能够使凝血原酶被转换成凝血酶，从而导致凝块形成。
APTT用作在外科手术过程中监测肝素治疗的常规测试、用作外科手术前的出血倾向的测试以及用于评价患者的凝固系统的综合能力。该测试通常是在中心实验室进行的。
激活凝固时间测试(ACT)
该测试类似于APTT测试并且用于在服用较大量的肝素的外科手术，诸如经皮经管冠状动脉成形术(PCTA)和心肺旁通手术过程中监测患者的凝固状况。ACT测试被认为是对于治疗血栓栓塞性疾病患者和进行体外循环治疗的患者肝素治疗控制的最好实验室测试之一。对于使用肝素的患者，ACT的延长与血液中的肝素浓度成正比。监测是重要的并且肝素剂量不足或者剂量过大可能会分别导致病理血栓形成或者严重的出血状况。
原始ACT测试使用带有C盐激活剂的玻璃管并且必须每15-30秒使装血的管倒置以使血液试样连续反复暴露在大量玻璃下。Helena实验室已经开发了MAX-ACTTM测试，该测试方法无需使管倒置，同时利用附加的玻璃珠使血液试样充分暴露在玻璃下。
凝血酶时间测试(TT)
该测试利用凝血酶对血纤维蛋白原的作用来测量血纤维蛋白凝块在血浆中的形成相对于正常血浆控制的比率。该测试通过在已经失去血小板的患者血浆中加入标准量的凝血酶并且测量凝块形成的时间来实现。该测试已经用于弥漫性血管内凝血和肝病诊断中，并且通常是在中心实验室内进行的。
其它测试
已经开发了针对特定因子的凝固化验，诸如表示因子IX不足的因子VIIIa。另一个实例是关于因子VIII的化验，这构成了血友病的测试。其它测试包括用于测量活性肽因子IXa、抗凝血酶、蛋白质C和蛋白质S的水平的化验。
已经开发免疫化学检验以鉴定和测量凝固和血栓形成的各个指标。
已经开发了用于实验室的各种仪器，诸如POC。此外，已经开发了能够让患者在家中对他们的血液凝固进行监测的装置。这特别适用于接受长期抗凝血剂治疗的患者，诸如丙酮苄羟香豆素。
下面举例说明用于测量血液凝固的各种技术。
转让于International Technidyne Corporation的US5,534,226披露了一种对血液试样进行凝固时间测试的设备和方法，其中利用设置在一次性使用的试管内的储液容器使血液沉积在毛细管中。接着使试样在毛细管内往复移动并且使血液被转移到一个限制区域。当转移限制区域所需时间比以前的时间长预定的百分比时，确定凝固已经发生。
转让于Hemosense的US6,060,323披露了一种用于血液试样的凝固或者消散的测量的一次性使用的电子装置和测试卡。使试样与两个电极接触，所述电极测量对应于试样凝固时的粘度变化的阻抗变化。
转让于Cardiovascular Diagnostics的US4,849,340披露了一种用于确定凝血酶原时间的光学检测方法，其中利用毛细作用将预定体积的液体试样抽到反应室中。使磁性颗粒与反应室中的试样混合，接着利用振动磁场进行搅动。光照射在试样上，接着进行检测。根据磁性颗粒移动程度的变化确定凝固点。
WO96/00390披露了一种用于确定血凝活性的完全抛弃型一次性使用的装置，其中利用试样沿着一种多孔基体移动的距离表示凝固时间。
转让于Biotrack的US5,039,617披露了一种用于对毛细管血液试样进行激活部分凝血酶原时间(APTT)分析测量的方法和毛细管流动装置。利用毛细管中的血流停止测量凝固时间。可利用流量或者压力传感器确定流速。毛细管的宽度可在0.05-3毫米的范围内并且需要不大于40微升的试样体积。或者，如果试样包含颗粒，那么可通过观察由于光源(例如LED或者激光)与毛细轨中的搅动颗粒相互作用所产生的散斑图可检测流量。
已经对凝固血样的变化阻抗之间的关系进行了研究(AmericanJournal of Clinical Pathology 67：470-476，1977)。对血样阻抗随时间变化的进行测量，所得到的阻抗曲线表示在凝固过程中所包含的各个步骤。
血小板凝聚的测量
血小板是直径为2-4微米的无色细胞片段并且存在于血液中。正常的血小板数量在180,000-400,000/微升的范围内，但是50,000/微升的血小板数量对于正常止血是足够的。在血管损伤后，例如在手术后，需要较高的血小板数量，有时超过100,000/微升。血小板的作用是通过自身粘附或者粘附于受损组织来修补血管壁中的裂口。当细胞受损时，它们释放某些能够使血小板从圆盘状变为球形并且变粘的化学物质，被称为凝聚粘附反应。人们已经认识到，血小板被在缺血性心脏病的病理发生中起到重要作用；急性心肌梗塞和不稳定心绞痛是与某些血小板因子的浓度增大相关的临床状态。另外，血小板功能失调是导致在心肺分流术后出血的几个重要的原因之一。人们还已经认识到，血小板通过释放增长因子有助于动脉粥样化形成的长时间过程并且血小板的功能还会受到高和低密度的脂蛋白的影响。这样，血小板功能的监测是一项重要和常规血液测试。
通常，利用Born血小板凝集计对分别标记为PRP和PPP的富含血小板和血小板贫瘠的血浆试样进行该测量，Born血小板凝集计能够测量透过试样的光传输。
US4,319,194披露了一种能够对全血进行血小板分析的血小板凝集计。金属丝形电极被插入到其中加有凝集剂的血样中并且阻抗的变化作为时间的函数被记录。但是，金属丝的移动导致电极间距离和阻抗测量的变化。
转让于Chrono-Log Corporation的US6,004,818披露了一种用于测量血小板凝集的方法，其中使试样在形成沟道的电极顶端的相邻平行表面之间流动。电极与用于搅动试样的装置一起被放入到充有血样的试管中。
发明内容
本发明的一个目的是，提供一种简单和廉价的能够对全血或者血浆试样(这里被定义为试样流体)进行凝固时间测量的设备和方法。
本发明的一个方面提供一种包含至少一个微通道的一次性测试装置，所述一次性测试装置能够被插入到用于确定试样流体的凝固时间的仪器中。
本发明的另一个方面提供一种集成的穿透装置和微通道以使微通道与穿透装置流体连通。
本发明的另一个方面提供一种用于测量这里所述的微通道内的试样流体的凝固时间的装置，以及利用沿着微通道的内和/或外表面设置的电极确定在使用状态下的在所述流体沿着所述微通道的流动已经停止时间或者位置的装置。
本发明的另一个方面提供一种用于测量试样流体的凝固时间的方法和装置，其中使试样流入到微通道中，从而流体阻抗变化作为时间的函数被监测。
具体而言，本发明的一个方面提供一种用于测量流体的凝固时间的装置，其包括用于采集流体试样的输入口，和至少一个具有横向尺寸小于约500微米的微通道，所述输入口与所述至少一个微通道流体连通，并且，至少两个电极沿着微通道的长度设置，其中，所述至少一个微通道被构造成利用毛细作用将流体样品沿其中传送，并且所述输入口是一整体式的刺入装置，其适于刺入使用者皮肤中以采集毛细流体试样，所述刺入装置的刺入尖端具有允许流体样品流通的通道。
本发明的另一方面提供一种用于制造测量流体的凝固时间的装置的方法，所述装置包括用于采集流体试样的输入口，和至少一个具有横向尺寸小于约500微米的微通道，所述输入口与所述至少一个微通道流体连通，并且，至少两个电极沿着微通道的长度设置，其中，所述至少一个微通道被构造成通过毛细作用将流体样品沿其中传送，所述方法包括：提供第一叠层；对所述叠层进行微制造以产生微通道、流体输入口和可选择的流体采集容器；沿着所述微通道的长度设置电极；提供带有电极的第二叠层，所述电极设置在其表面上；以及层压所述第一和第二叠层。
本发明提供一种一次性测试装置，所述一次性测试装置包括支撑件，至少一个微通道设置在所述支撑件上或者内。微通道的作用是接收和容纳流体试样。由微通道接收的试样与存在于通道内的凝固激活因子混合，所述凝固激活因子最好涂在微通道和/或试样采集容器的内表面上。接着，试样利用毛细作用沿着通道长度流动直至其停止流动或者已经确定流量小于某一阈值。然后，将通过对位于通道内或者位于通道外的电极之间的电容或者阻抗的测量间接确定所移动的距离。在沿着微通道的任何适合的位置处设置排气孔以能够使试样流动，接着排出内含的空气或者气体。
可设置与所述一个或者多个微通道流通的容器以使流体试样可容易被采集。或者，可利用输入口将试样直接输送到微通道。
本发明的微通道沿着其长度可具有恒定的直径或者变化的直径。由于试样和壁之间的接触面积增大而导致摩擦增大，因此沿着毛细管长度的流率会随着通道充填而减小。在毛细管中实现恒定流率的一种方法是如在US4,756,884中披露的，直径作为毛细管长度的函数被增大。
但是，对于一些测量，流率随着长度的降低可能被认为是有利的。通常，在凝固被认为已经发生之前，APTT的测量需要花费高达500秒的时间。在这样长的时间内保持流动需要长的通道长度，因此任何的流率降低将用于减少所需的总试样体积。
可设置与微通道流体连通的并且体积大于微通道的容器。这样一个腔室的作用是最初采集试样。在使用过程中，试样被供给到输入口，试样在输入口处被抽到腔室中。例如，如果试样是通过切割手指获得的毛细管血，那么使用者能够在腔室已经被充填到适合的量后移开手指。试样接着进入和沿着较小直径的毛细通道移动。凝固促进因子诸如促凝血酶原激酶可被涂在腔室的内壁上或者涂在毛细管的壁上。
如上所述，设置一个或者多个排气孔以能够使流体沿着毛细管流动。排气孔可用于控制流体流动。例如，可设置第一排气孔以使流体进入所述腔室但没有进入到随后的毛细通道中。接着可打开第二排气孔以使流体沿着微通道通过。或者，可利用其它流动控制装置控制流体的流动。任何适合的流动控制方法可与相应的装置结合使用以实现这样一种控制方法。例如，压电泵、电动或者机械方法诸如解除沿着所选择的导管的流动堵塞-例如通过使气泡逸出或者通过打开一个阀。在某些实施例中，流动控制装置包括位于导管/微通道内的疏水性门。这里所述的疏水性门指的是在亲水性通道内的疏水性表面区域，以使流体流动被中断。通过改变门的疏水性，即，增加疏水性区域的亲水性，接着可使流体沿着通道流动。可使用疏水性门控制流体在单个微通道内的流动或者可使用疏水性门将流动从一个微通道切换或者改向到另一个微通道。
或者，可保持该门的疏水性并且可施加较大的泵送力(例如由机械或者电渗泵提供的)以使流体突破疏水性门。
沿着微通道的长度设置能够提供试样电性能测量基础的电极，并且电极可沿着其整个长度或者沿着其长度的一部分延伸。电极还与支撑件上的接触点电接触。该条被设计成被插入到一个仪器中以使支撑件上的接触点与该仪器中的相应的接触点接合和电接触。接着，将由电极测量的电参数传送到能够翻译信号的仪器以提供结果。该仪器还将存储校准信息以能够提供INR值。
电极可是任何适合的惰性导体并且特别可从碳、金或者铂中选择。电极可沿着微通道的整个长度或者其一部分延伸。可通过在微通道上印刷油墨或者其它沉积方法，例如真空沉积或者溅射来生产电极。电极可沿着通道的外部和内部延伸。电极位于微通道外部的一个优点是，可通过其实际形状和化学组分干扰凝固过程的电极不与试样接触。另外，碳能够吸收血液中的物质，从而影响表面化学性质。
电极可采用任何适合的形状和尺寸并且通常采用在任何位置处的宽度可在通道周长的1-99％之间的线或者薄带形式。电极通常位于通道的相对两侧上并且不一定具有相同的宽度。如上所述，电极基本上覆盖了所有内表面积或者外表面积或者仅一小部分。根据一个实施例，通过形成充填有导电材料的第一通道和形成用于输送测试流体的第二通道将电极设置在流体通道的相对两侧，第二通道横截第一通道，从而在第二通道的各相对的两侧内形成两个导电部分。另外，在该装置具有利用电渗力工作的流动控制装置的情况下，驱动电极最好彼此相邻设置。这能够在无需提供不希望的高电压的情况下形成高电场。
根据本发明所形成的导电部分用于电化学传感器装置。用于制造上述相交通道的微加工技术是优选的，这是由于它们可用于制造彼此相邻设置的微通道，使其密集排列。
这里所用的术语“微通道”指的是一种具有任何适合的横截面并且其最小横向尺寸小于约500微米的通道。在本发明的优选实施例的微通道中，其尺寸小于200微米较好，最好在约10-200微米之间。通道的长度可取决于具体的测试形式的任何长度。但是，通道的长度可在1-10厘米之间并且提供200nl-2ul(微升)的体积。例如，直径为40um的30厘米通道的总体积约为19ul。
对于一种分析物感测装置，由于多个原因而使这样的微通道是有利的。进行化验所需的流体体积相应是小的。对于体液测量，小试样体积是有益的，这是由于这意味着比较容易提供足以用于进行有效测试的体积。该测试可利用从毛细血管例如手指刺入或者其它适合的切割点中获得的全血试样来进行。试样体积的减小还相应减小疼痛，这是由于可使用较小的针来刺破皮肤。
应该理解的是，将微通道结合在这样的测量装置中，可对凝固所用的时间进行精确地确定，这是由于即使对于凝固形成不良的试样，这里所述的微通道的较小横向尺寸意味着少量凝固阻止流体在其中流动。另外，微通道总体上将需要很小的试样体积并且无需牺牲其长度可盘在或者安装在合理的表面积上。通道长度的最大化是重要的以适应长的凝固时间-例如，对于血液的PT测量需要20-30秒并且提供较大的测量分辨率。
用于确定流动停止的时间的装置可直接使用，例如，它可包括流率传感器，最好是一种能够以电的方式检测这样的流率的传感器。显然，当检测的流率为零或者至少在一个阈值以下时，可以确定流动已经停止。
利用横穿通道的两个电极之间的阻抗变化率来测量流率。电极最好可采用前面所述的形式。可测量阻抗的电阻部分。例如，电极阵列可沿着通道间隔布置以根据血样已经移动的距离来提供一个增量信号。或者，可沿着通道壁设置单个大电极，它与反电极之间的电阻取决于大电极被覆盖的程度。
最好测量阻抗的纯电容部分。这意味着电极无需与试样流体接触。另外，一系列间隔的电极可提供不连续的读数，或者最好可形成单对细长电极-例如，在通道的相对两个壁上。应该理解的是，两个“板”之间的电容将取决于通道充填血液的程度，从而取决于空气和血液的电容率(介电常数)相对差。
对于光学感测技术，当横穿窄的毛细管进行测量时，由于很短的路径长度而会出现困难，路径长度取决于光学信号的强度。或者，平行于通道的照射光要求光学部件的精确对准以及使用镜子。另外，在凝固过程中，试样的前沿具有首先凝固趋势。因此平行于通道引导的光透过未凝固和凝固的血液，使测量变得较复杂。另外，例如由于光学信号散斑图，因此所产生的光信号是复杂的并且需要复杂算法来评价结果。光学系统也是昂贵的，需要光源和检测器以及存在将它们放置在何处来观察凝固过程的问题。对于本发明，不存在由于检测系统的放置而带来的这样的困难，这是由于电极沿着通道延伸。由于所测量的阻抗间接与微通道的直径成比例，因此从这一点出发，很小的直径实际上是有利的。电极沿着通道的长度设置，因此由电极测量的阻抗是一种累积测量，取决于由试样覆盖的长度或者体积。这样，关于将电极放置在何处以监测凝固过程的问题不是这样一个关键问题。另外，基于这种方法的监测系统与光学系统相比，在制造上更简单和廉价。
插入内容
平行板电容器的电容如下表示：
C＝ε0εrA/d
其中：
ε0＝自由空间的电容率
εr＝板之间的电介质的相对电容率
A＝板的表面积
d＝板之间的距离
假设电极具有恒定的宽度w和长度1，该等式变为
C＝ε0εrwl/d
现在，如果通道被血液部分充填的距离为x并且相对电容率为ε1，而通道的其余部分是空的，具有相对电容率ε2，那么通道的两个相邻部分可被认为是分开的电容器。
被充填的部分的电容是
Cfilled＝ε0ε1wx/d
空的部分的电容是
Cempty＝ε0ε2w(1-x)/d
由于这两个电容是并联的，因此总电容等于它们的和，即：
C＝ε0ε1wx/d+ε0ε2w(1-x)/d
＝ε0w/d(ε1x+ε21-ε2x)
通过测量总电容C并且知道其它常数，因此可计算血液移动的距离x：
C＝ε0w/d(ε21+x(ε1-ε2))
dC/ε0w＝(ε21+x(ε1-ε2))
x＝1/(ε1-ε2)(dC/ε0w-ε21)
从上面内容可以看出，还能够监测电容的变化率以确定流动停止所用的时间，能够测量血液在凝固前移动的距离x的数值。这提供了一种凝血酶原时间的相对测量，这是由于血液凝固所用的时间越长，其沿着通道前进的距离越长。
因此，这例如可用作直接时间测量的交叉检验。
从上面的等式中可以看出，电容，特别是通过将血液引入到板之间所达到的电容变化还与它们之间的距离d成反比。这样，可以看出，利用在与板正交的方向上的横截面尺寸远小于与板平行的方向上的横截面尺寸的通道可以实现电容C数值中的较高的绝对变化。尽管提供了大的电容变化，但是这可能与在凝块形成时快速阻止流动的需要矛盾。这样，在另一个实施例中，提供一个微通道的平行阵列，每一个微通道带有一对电极，并且测量电容中的累积变化。这可提供同样大的变化，但不会损害流动被凝块形成阻止的倾向。对于其中带有电极的微通道，除了具有上述应用以外，还有许多其它的可展望的应用，因此在较宽的方面上，本发明提供一种包括微通道和其中的一对电极的装置。在另一个实施例中，可提供微通道阵列，每一个具有不同的直径。多个通道或者单个通道可具有恒定的直径，或者沿着其长度可变的直径。例如，通道的直径可被改变以使试样流动加快或者减慢，这样能够使其有效地溶解位于通道表面上的凝固促进化学物质。不同管内的不同的凝固促进化学物质可被提供作为表面涂层，这样例如可同时进行PT和APTT的测量。
微通道本身可由具有适合物理性质的材料制成。微通道应该具有良好的导热性、平滑的毛细流动，为试剂提供均匀的涂层，以及本身不会促进凝固过程。该材料还应该确保在凝血已经发生后，流动应该停止或者减慢。例如，研究已经表明，与硼硅酸盐或者商业上的硅化硼硅酸盐接触可大大缩短PT。因此应该避免使用这样的材料。如果需要的话，该通道也可涂有毛细阻滞剂或者促进剂。
在另一个实施例中，微通道可被这样设计，即，使试样流入微通道中，充填通道至预定深度后横向流动停止。在这种情况下，电极可被设置在通道的内表面或者外表面上，电极沿着通道的整个部分或者一部分延伸。另外，通过测量阻抗随时间的变化可以观察凝固过程。通过对阻抗曲线的测量和后续分析，人们可以确定凝固的开始。
可利用任何适合的技术制造本发明所涉及的微通道。特别是，如果允许的话，可利用任何适合的微制造技术，诸如压花、等离子蚀刻或者激光照相烧蚀制造微通道。对于微通道的材料，可利用任何适合的微制造塑料，诸如聚酯、聚碳酸酯、聚苯乙烯或者聚酰亚胺制造微通道。优选的聚合物是聚碳酸酯。这些允许利用后续的激光修整产生附属的微或者纳结构(例如，任何所需的图形或者其它修整可形成在微通道中)。聚苯乙烯在叠置过程中表现了更好的性能。这样，聚碳酸酯可用作下叠层，聚苯乙烯可用作上叠层。正常的叠置方法使用涂有压敏或者热熔粘结剂的薄片将一个薄片结合在基体上或者另一个薄片上。这样一个标准方法可能存在与上述装置相关的问题。首先，需要适合于印刷方法的薄片。由于印刷设备内的问题而使这对于任何压敏粘结剂是困难的。可利用涂有热熔粘结剂的薄片来解决该问题，其中粘结剂仅在高温下(例如80℃)变粘。利用该系统使得用于印刷电极和其它结构的油墨沉积变得很容易，但它在叠置步骤中存在问题。胶层在高温下基本上变成液体，因此印刷结构失去形状并且受到拉伸和出现变形。这样的变形不仅是电极的装饰问题，它改变电极表面(其与响应信号成正比)以及材料的内电阻和电催化性能。
除了上述问题以外，存在胶进入通道和使通道凝固或者变形的附加问题。对于上述芯片，使预印刷的薄片与芯片基板结合的最优选的方法是无粘结剂热粘合方法。利用冲压工具或者热辊压力机在高温下进行粘结。该温度接近聚合物的玻璃态转变温度(Tg)，这样该聚合物的低分子重量部分变得可动和变粘，而该聚合物的高分子重量部分仍然支撑薄片或者薄膜的整体。聚合物的低分子重量部分将两片粘合在一起(基板和带有电极的薄片)，另外它将符合印刷的电极的形状，厚度可在5和30微米之间。因此，人们不会看到基板和印刷区域之间的泄漏。利用相同的热塑性聚合物实现理想的粘合，诸如聚苯乙烯在聚苯乙烯上或者聚碳酸酯在聚碳酸酯上。但是，在适合的状态和温度/压力下，结合的聚碳酸酯也可粘合在聚苯乙烯上。但是，硬质塑料(非热塑性)材料不适于这样一种方法。
根据各个参数的适用性，诸如其微制造能力、对于凝固过程的惰性、亲水性、形成光滑通道的能力、其热容和导热性、在其表面上携带电极的能力、坚固性等选择材料。如果需要的话，该材料还可具有影响亲水性从而影响毛细性能的表面涂层。这又确定血样的流率。
可利用任何适合的技术制造本发明所涉及的测量装置。特别是，如果允许的话，利用任何适合的微制造技术制造微通道，诸如(但不限于)压花、等离子蚀刻或者注射模制。
一个或者多个电极可形成在第二基体上，第二基体接着叠置在该装置的主支撑件上。用于将电极沉积在基体上的方法可从印刷方法中选择较好，最好采用一种丝网印刷方法。或者，也可使用化学或物理气相淀积技术。通常说来，本发明所有实施例所涉及的电极都可用任何合适的惰性物质(诸如碳、金、铂等)构成。依照一个实施例，使用丝网印刷可将选择性地涂覆有试剂的碳电极设在第二基体上，而后所述第二基体被叠置在支撑件上从而封闭该通道或多个通道。这提供了一种适用于电极形成在封闭通道内的实施例的很简单的制造方法。
通常这样执行一个基体到另一个基体上的叠置，即，使得两层完全对齐，并且无需修整或剪切。然而，也可如下制造该装置，例如，通过首先是叠置步骤，其后是剪切步骤，从而可将第二基体修整为支撑件的形状。可通过各种方法执行叠置，诸如超声波、或热焊接或粘合，或者使用粘合剂。在将上基体层叠置在支撑件上之前，微通道和/或容器的壁可被涂有一层凝固促进剂，诸如由DADE Behring提供的Thromborel R(商标)促凝血酶原激酶凝固剂。
该带还可包含用于对支撑件加热的装置，以使血液或者血浆可在微通道内被加热到预定的温度。这可被设置在支撑件自身上或者微通道的外表面上。这样的装置可采用与仪器电接触的附加电极的形式，以利用从仪器流到电极的电流来产生热。或者，通过将带插入到仪器中来实现对试样的加热，热量可由仪器本身产生，可利用对流使试样变温。另外，可提供温度控制装置以对加热装置的接通或者断开的切换或者加热速度的改变进行控制。这样可使试样保持恒定的温度或者特定温度范围内。最好将加热装置设置在带自身上，这是由于可使试样更快被加热并且能够更有效地对温度进行控制。根据一个实施例，测量电极也可用作加热电极。
该带也可具有用于检测已经提供足够的试样的时间从而开始进行测量的装置。这样的充填检测装置可被设置在流体容器内并且可包括两个分隔的电极，在两个分隔的电极之间的测量可确定容器的充填状态。存在于所述仪器中的控制单元可控制任何流动控制装置的工作并且能够显示该装置的任何故障。由充填检测电极确定为满容器的指示将向控制单元发送一个信号，控制单元接着将控制任何流动工作装置以使试样沿着毛细管流动。
该仪器能够显示测试的结果。另外，该仪器具有记忆功能，能够下载信息以及具有下载于其上的信息。该仪器也可装有一种算法以便在需要的情况下指示根据结果所需要采取的动作。该仪器可是一种能够下载和接收医生或者网站的信息的无线通信装置。该仪器还具有存储和翻译关于患者的个人信息的能力。该仪器还具有远程阅读关于该带的信息的能力以及能够远程存储批校验码，即光学等。
依照一个实施例，支撑件在其一端形成有整体针，而后第二基体被叠置于支撑件上形成一个通道，并使得刺入元件露出。依照另一个实施例，整体皮肤刺入元件被设置在一端上的开口处。刺入元件被如此布置，即，使得上部被插入到皮肤中，而皮肤本身有效地形成元件的壁，从而它可用作是空心针。更好的是，通过形成这样的刺入元件而实现这一点，所述刺入元件具有远离开口侧逐渐变细的壁-例如，V形。因此从本发明的另一个方面来看，本发明提供了一种用于获得并测量流体的设备，所述设备包括具有至少一个纵向侧开口的皮肤刺入元件，另一侧可被如此布置，以使得当将刺入元件插入到皮肤中时被刺入的皮肤用于保留元件的纵向侧。
因此，从本发明的另一个方面来看，本发明提供了一种制造用于测量流体中分析物浓度的装置的方法，所述方法包括以下步骤：提供支撑件、在支撑件的表面上形成开放式通道以及在所述支撑件上叠置第二层以便于封闭所述通道。本发明还扩展为使用这样一种方法制成的装置。
所述针最好适于刺入所述皮肤。例如所述针的尖端区域最好是大致圆锥形。另外，所述尖端区域具有较小的横截面，宽度小于0.2毫米较好，最好宽度小于0.05毫米。
另外，所述针最好能够在插入皮肤后使其被堵塞的危险达到最小。例如，针孔可设置在针的侧面上，而不是如常规的设置在尖端处。针孔最好是凹入的，从而在刺入后避免与皮肤接触，从而避免可能出现的堵塞和/或受损。
所述针最好具有能够使试样流体在毛细作用力下被向上抽吸的孔。
该装置适用于测量血液或者血浆中的凝固时间。尽管该装置适于测量凝血酶原时间(PT)，但是利用该技术可测量其它凝固时间，诸如激活部分凝血酶原时间(APTT)、激活凝固时间(ACT)和凝血酶凝固测试时间(TCT)。
附图说明
下面将结合附图，仅通过示例的方式，描述本发明的一些优选实施例，在附图中：
图1和图2示意性地示出了一种适于测量血液凝固的装置；
图3示意性地示出了一种带有螺旋微通道的适于测量血液凝固的装置；
图4(a)示出了包括多个通道的本发明所涉及的带；
图4(b)示出了图2中的可选择的实施例，其中通道具有不同的横截面尺寸；
图5a示出了一种整体式刺入装置和微通道；
图5b-5h示出了刺入装置的其它视图；以及
图6(a)和图6(b)示出了另一种可选择的结构，其中微通道被一系列纵向间隔的成对的电极侧面连接。
具体实施方式
如图1中所示，其中提供包括上支撑件101和下支撑件103的一次性测试带100。微通道102形成在下支撑件103的上表面中。第二基层101叠置在支撑件103的顶部上，从而封闭开放的微通道102。
电极104形成在基层的相应的外表面上并且与通道共面。如图1a中所示，电极可沿着整个外部长度延伸到该装置的一个边缘，以使导线和仪器(未示出)之间形成适合的电接触。电极自身可被另一个叠层覆盖以对它们进行保护。图中还示出了容器105和试样输入口106，试样最初流到容器105中。如图中所示，输入口基本上与微通道平齐并且利用毛细作用充填容器。但是，试样输入口可设置在该装置的上方并且可利用重力作用充填试样容器。或者，容器可设置在该装置的外部以使其还用作一个试样采集装置。容器可安装在带的顶表面上并且可利用任何常规的方法形成，诸如注射模制。图1中未示出能够使空气从通道中排出的排气孔和加热或者充填检测电极或者流动控制装置。流动控制装置可沿着微通道自身设置。这些图仅是说明性的，并且没有反映该装置内的部件的相对尺寸。这样，容器相对于微通道具有特定的体积以使其可存储足够体积的试样，从而可进行测量。所示的容器的形状为矩形，但是它也可采用任何适合的形状。
图2示出了另一个实施例，其中电极位于叠层的内表面上。或者，可沿着通道的内部或者外部以一定间隔设置电极。作为图1和图2中的电极布置的另一种可选择的方式，电极可设置在上叠层或者下叠层上。
图3中示意性地示出了另一个可选择的实施例。可以看出，在该实施例中，微通道202采用螺旋状以对于装置的已知的表面积可增加长度。
本领域普通技术人员应该理解的是，两个电极之间的电容特别是与微通道中的内含物的相对电容率成比例的。假设血液的相对电容率与水基本相同，因此约为80的数量级。另一方面，空气的相对电容率约为1。
因此，两个电极104之间的总电容取决于充填有血液的微通道102的比例。因此可通过测量电容能够获得血液的凝血酶原时间的另一种可选择的评价方式。如果两个不相容，那么这相对于凝血酶原时间在经验上进行校准并且用于指示错误需要进行重复测量。
相对于螺旋形微通道结构，示出了与微通道流体连通的输入口106，且可用于任何结构。输入口是这样设计的，即，使接触该输入口的试样能够被输送到微通道中。所示的前沿109为平的。但是，它可采用圆形或者其它任何适合的具有人机工程学优点的形状。或者，试样可被供给到与微通道流体连通的容器中。
图4a示出了另一个可选择的实施例，包括多个通道。这些通道具有公共的输入口，但是所述公共的输入口是这样设计的，即，使流体不能相互流通。图4b示出了具有不同尺寸的多个通道。选择不同的尺寸，能够改变横截面积与接触通道壁的血液的周长的比率。除了具有多个直径不同的通道以外，一个或者多个通道可沿着它们的长度具有改变的横截面积。
图5a-5c示出了本发明的另外两个实施例，其中示出了整体式刺入元件和适于测量流体凝固的微通道。参见图5a中所示的装置115，可以看出，它基本上由层116制成的，第二层(未示出)附于或者叠置在层116上。最靠下的基层116包括模制的或者冲压的微通道118，以及整体形成的刺血针119与微通道的入口103相邻设置。在制造过程中，微通道118可涂有适合的试剂，诸如凝血酶原，可利用任何常规的装置来提供，例如诸如丝网印刷或者喷墨印刷的印刷，或者在制造过程中喷涂。最上层117附于下表面上以使测量电极位于所形成的通道的下侧。或者，电极可位于叠层117的外表面上。图5a-c未示出用于采集流体的容器、充填检测电极、排气孔或者加热电极。一个或者多个排气孔可设置在任何常规的位置处。图5a也没有示出另一个电极，在它们之间进行电参数的测量，即，阻抗或者电容。在电极的端部和位于测量仪器内的适合的连接点之间进行电连接。上层117可略长于下层116以能够接近轨321。根据该特定实施例，沿着上叠层117以一定间隔提供电极。或者，提供一个基本上平行于微通道的电极。图5a没有示出与前面描述的一致的可设置在下叠层116或者上叠层117上的任何适合位置处的“反”电极，即，在微通道的内表面或者外表面上。
应该注意的是，特别是在图5a中所示的带115的刺血针119的横截面基本上为V字形并且朝向其尖端逐渐缩小。这意味着，当利用其刺破使用者的皮肤123时，如图5b中所示，V字的两侧边回压皮肤123的一部分，迫使表皮形成被封闭的通道124的保持壁123。这样，当其被插入到皮肤中时，一个开放的通道被有效地转变为封闭的通道。这能够使流体被向上抽到所形成的通道124并且进入到微通道118中，无需模制一个很细的空心针。微通道118还形成有便于制造的V形型面，但是从图5c和图5d所示的略微改变的实施例(其中的微通道118′是矩形型面)中可以看出，这不是实质性的。
在带115的使用中，使用者首先将测试装置插入到仪器中。或者，该测试装置已经以单个装置或者多个分别组装在盒体内的装置的形式被装入一个整体式测量和刺入装置中。接着，使用者利用刺血针119刺破他们的皮肤，利用毛细作用使试样通过由刺血针119和皮肤122所形成的通道124流入到微通道118中，最好经过具有充填检测装置的充填容器以使试样开始流入到微通道中。
图5d至图5g示出了用于刺入皮肤的体液满载层的刺血针的可选择的实施例，作为图8a和图8b的刺血针119的实施例的一种替代。在图5d中，刺血针119a是整体形成的从装置115(在图5d中未示出，但是该装置115与图5a中的一样)处突出的尖锐突出部分，其中纵向毛细通道121a完全切入刺血针119a的厚度。在刺血针119a的尖锐远端125a处，刺血针119a具有通道121a的增大区域123a。该增大区域123a也完全切入刺血针119a的厚度。在其近端127a处，毛细通道与图5a的装置115的微通道118相连接。
如从图5g中清楚看出的，图5d的实施例允许流体从刺血针119a的相对侧进入到毛细通道121a，并且其中皮肤壁与刺血针119a的壁合作以限定出封闭的通道121a。然后流体可积聚在集合区域123a中并且从集合区域123a中流入到毛细通道121a中以及从皮肤直接流入到毛细通道121a中以便于通到微通道118中。
在图5e的实施例中，示出了与图5d的刺血针相似的刺血针119b的设计，但是图5e的刺血针119b不包括大的集合区域123a。集合区域123a的去除可允许通向刺血针119b的更窄的横向尺寸。除使用模制件制造所述装置以外，还可用导电材料冲压成基底元件116和刺血针119、119a、119b。在此类情况中，基底元件可为电极。可如上所述用金属冲压成导电性的基底元件和刺血针，或者可用任何其它可接受的方式(例如，光化学蚀刻金属原料，机加工或者其它制造技术)。虽然可用不锈钢制成导电性的基底元件，但是也可为其镀上第二层金属(比如金、铂或银)或者涂有介电绝缘体。
图5h示出了整体形成的基底元件和刺血针，所述整体形成的基底元件和刺血针最好是用一片金属薄板冲压而成的。所述金属最好为(但不局限于)可选择地涂覆有贵金属(诸如金或银)的不锈钢。在还示出了在底片上的微通道，其中第二层(诸如测试带)可被附于所述微通道上。图5h中还示出了具有矩形排气口81的冲压的刺入元件，所述矩形出口81还用作一种毛细管堵塞，所述毛细管堵塞可确保，一旦刺血针83将流体吸收到敏感区82中以后，流体流动就会被阻断。所述出口可为任何适合的尺寸或形状的。
图6a和图6b示出了另一种可选择的结构，其中微通道300被整体形成在微通道300的壁中的一系列纵向间隔的电极对38侧面连接。为了形成这些电极38，首先在基体材料302中切割一系列平行通道34。接着，通道34充填有碳使其具有导电性。然后，以与平行通道34直角的形式形成微通道300以使其与它们相交。这样在微通道300的每一侧上形成相对的电极38。
这种结构通过测量相邻电极对38之间的电阻来监测沿着微通道300前进的血液。当血液达到每一个接连的对时，电阻从断路下降至大约200千欧。这样，获得关于移动距离的间断读数。该结构还证实了，可使电极38与血液接触。与微通道流体连通的输入口206设置在支撑件的外表面上。排气孔207位于微通道的远端处以使空气或者其它气体可从通道中排出，以使试样进入。所述排气孔可设置在沿着微通道的任何适当位置处或者排气孔可设置在未密封的毛细管的端部208处。一组金属带或者丝209可沿着微通道的任何一侧设置并且沿着其长度延伸。这些金属丝本身与用于与仪器相连的一组触点210电接触。这些触点也可用于通过沿着导线提供的电流在插入试样之前对试样或者微通道进行加热。或者，可利用支撑件内或者上的导电带对所述带进行加热，如图1b中所示。
图1c示出了另一个可选择的实施例，其中微通道205设置在支撑件200上。
作为入口206的一个可选择的实施例，容器可用于采集流体试样。如图中所示，在图3中，容器位于带的顶侧并且用于与微通道流体连通。
根据本发明的一种方法，首先将测试带插入仪器中以使接触点形成与设置在仪器内的相应触点电连接。接着将血样供给到微通道102的端部，血样然后沿着微通道流动，此时定时器开始计时并且开始进行测量。或者，最初将试样经试样输入口抽到容器中，从而充填检测电极将确定足够的试样是否已经被提供。接着，流动控制装置使试样在毛细作用下被抽入到和沿着微通道移动。利用充填检测装置启动流动控制装置以使该装置无需进一步从使用者输入。充填检测装置也可用于接通该装置。当血液沿着微通道流动时，其与凝固剂接触使其凝固。这最后阻止血液沿着通道中途流动。两个电极101和102通过在带边缘处的连接部分106与测量电路(未示出)相连。该电路用于测量两个电极之间的电容。这可利用本领域已知的任何方式来实现，例如通过使该装置成为RC振荡器的一部分和测量其频率(与电容成反比)。当血液流动时，两个电极104之间的电容将改变。当流体停止流动或者当流率降至一个预定值内时进行测量。
利用标准化因子除以所测量的实际凝血酶原时间，正常血液凝固所需的时间与通道的尺寸和凝固剂的性能相关。采用国际标准化定量(INR)或者凝血酶原时间的数值形式的结果被显示在读数器上(未示出)。
本领域普通技术人员应该理解的是，虽然在文中更详细地描述了本发明原理下的某些可能实施例，但是对于这些可能性来说还存在许多不同的变化和改进。例如，本发明所涉及的装置可测量除血液以外的其它流体的凝固时间。
Claims (9)
1.一种用于测量流体的凝固时间的装置，其包括用于采集流体试样的输入口，和至少一个具有横向尺寸小于约500微米的微通道，所述输入口与所述至少一个微通道流体连通，并且，至少两个电极沿着微通道的长度设置，其中，所述至少一个微通道被构造成利用毛细作用将流体样品沿其中传送，并且所述输入口是一整体式的刺入装置，其适于刺入使用者皮肤中以采集毛细流体试样，所述刺入装置的刺入尖端具有允许流体样品流通的通道。
2.如权利要求1所述的装置，其特征在于，所述电极具有不同的长度和尺寸。
3.如权利要求1所述的装置，其特征在于，所述电极位于所述微通道的内表面上。
4.如权利要求1所述的装置，其特征在于，所述电极位于所述微通道的外表面上。
5.如权利要求1所述的装置，其特征在于，用于促使流体凝固的试剂设置在该装置的内表面上。
6.如权利要求1所述的装置，其特征在于，其包括与用于采集流体试样的通道流体连通的容器。
7.如权利要求1所述的装置，其特征在于，其包括多个微通道。
8.如权利要求1所述的装置，其特征在于，所述微通道的横截面积沿着其长度改变。
9.如权利要求7所述的装置，其特征在于，所述微通道具有不同的直径。
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| GB0030929.4 | 2000-12-19 | ||
| GBGB0030929.4A GB0030929D0 (en) | 2000-12-19 | 2000-12-19 | Analyte measurement |
Publications (2)
| Publication Number | Publication Date |
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| CN1491358A CN1491358A (zh) | 2004-04-21 |
| CN100458441C true CN100458441C (zh) | 2009-02-04 |
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| Application Number | Title | Priority Date | Filing Date |
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| CNB018227287A Expired - Fee Related CN100458441C (zh) | 2000-12-19 | 2001-12-19 | 用于测量血液凝固的装置及其方法 |
| CNB018227333A Expired - Fee Related CN1292703C (zh) | 2000-12-19 | 2001-12-19 | 分析物测量 |
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| Application Number | Title | Priority Date | Filing Date |
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| CNB018227333A Expired - Fee Related CN1292703C (zh) | 2000-12-19 | 2001-12-19 | 分析物测量 |
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2001
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2004
- 2004-10-20 HK HK04108215.8A patent/HK1065591A1/xx not_active IP Right Cessation
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