CN1296963C - 用于检测和识别空气中的生物气溶胶颗粒的方法和装置 - Google Patents
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Abstract
在用于检测和识别空气中的生物气溶胶颗粒的方法中，通过荧光技术在ATOFMS(气溶胶微粒飞行时间质谱分析仪)中选择颗粒流中的生物气溶胶颗粒，且仅使所选的生物气溶胶颗粒电离，例如在MALDI(基质辅助的激光解吸/电离)基础上，此后检测生成的离子，并识别出生物气溶胶颗粒。生物气溶胶颗粒的选择是通过激光辐射进行的，所述激光由第一激光装置产生，其波长在生物气溶胶颗粒的特定物质中产生荧光，此后，通过荧光检测器选择生物气溶胶颗粒，然后触发第二激光装置发射光束，其波长使仅由荧光检测器选择的生物气溶胶颗粒电离。
Description
技术领域
本发明涉及一种用于检测和识别空气中的生物气溶胶颗粒的方法和装置。
背景技术
这种方法及其所使用的测试装置可以从M.A.Stowers等的论文：基质辅助的激光解吸/电离对在线气溶胶微粒飞行时间质谱分析的应用(Application of matrix-assisted laser desorption/ionization to on-line aerosoltime-of-flight mass spectrometry)，Rapid Commun.Mass Spectrom.14，829-833(2000)中可知。该文指出，对于“准实时生物气溶胶微粒检测”有两种选择，即利用生物分子的固有荧光性的方法、和将质谱分析应用于激光解吸产生的离子上的方法。因为生物气溶胶颗粒仅形成空气中的气溶胶颗粒总量的一小部分，所以可以使用公知的“单颗粒”荧光检测器，指示气溶胶颗粒是否是生物起源或非生物起源；然而，它们不能识别生物气溶胶颗粒。所以，上述论文描述了一种识别试验，该试验从雾化液体中的生物气溶胶颗粒开始，然后将喷雾吸入ATOFMS(气溶胶微粒飞行时间质谱分析仪)中。在ATOFMS中，在MALDI(基质辅助的激光解吸/电离)(matrix-assisted laser desorption/ionization)的基础上进行气溶胶颗粒的识别。对于这一试验，即使首先制备了气溶胶微粒，但该论文中示出的研究目的仍然针对实时基础上进行的识别。然而，按所述方式，不能令人满意地实现，因为空气中大部分含有非生物气溶胶颗粒。因此，所使用的ATOFMS检测和识别设备对生物气溶胶颗粒不很灵敏。考虑到在特定情形例如在生物战的情形下生物气溶胶颗粒的快速检测和识别是绝对需要的事实，本发明的目的是针对上述论文中陈述的目标，即快速的在线生物气溶胶颗粒检测和识别。
快速的在线气溶胶颗粒检测和识别，不涉及生物气溶胶颗粒的检测和识别，在这方面，已经可从国际专利申请WO96/31900中得知。根据该申请，含有颗粒的气流经过ATOFMS系统。以这种方式，快速检测和识别特定的生物气溶胶颗粒是不可能的，因为，如前所述，空气中的生物气溶胶颗粒较少，且它们一在空气中散布，就必须精确地检测和识别。
发明内容
为了解决上述问题，在本发明的方法中，在ATOFMS中通过荧光技术选择颗粒流中的生物气溶胶颗粒，随后使选择的生物气溶胶颗粒电离，检测所生成的离子，从而识别生物气溶胶颗粒。对于生物气溶胶颗粒的选择，利用本身公知的特性，即，当用适当的波长照射时，特定物质比如氨基酸的存在导致产生特有的荧光。因此，通过266nm的UV激光照射色氨酸呈现出在300至400nm的波长范围的较宽荧光。而且，合格的是比如酪氨酸、NADH或核黄素的物质的荧光光谱(参见：Fell等：浓度、大小和激励功率作用和含有色氨酸和细菌的微粒，SPIE，vol.3533，pp.52-62)。总之，无机和大多数有机物质不表现出这一特性。为了在本发明的方法中利用这一特性，通过激光辐射进行生物气溶胶颗粒的选择，所述激光通过第一激光装置产生，其波长在生物气溶胶颗粒的特定物质中产生荧光，此后，通过用于检测荧光辐射的检测器选择生物气溶胶颗粒，然后触发第二激光装置发射光束，其波长使仅由荧光检测器选择的生物气溶胶颗粒电离。对于第一激光装置，最好采用连续波激光装置(cw激光装置)，而对于第二激光装置，使用脉冲激光装置，所述脉冲由荧光检测器触发。
在选择生物气溶胶颗粒过程中利用气溶胶颗粒的大小也是有效的，即空气动力学尺寸。细菌和病毒的大小基本上在20μm以下的范围内。因为气溶胶颗粒以给定的速度进入ATOFMS的中央空间中，从气溶胶颗粒经过已知距离的持续时间可以确定连续的气溶胶颗粒的大小。通过使第一激光装置的激光束照射在两个具有已知相互距离的连续点上，上述持续时间因而也就是气溶胶颗粒的大小可以从气溶胶颗粒散射和检测的光确定。然而，所述点之间的距离必须小于连续颗粒的相互距离。例如，在所述点之间的距离为2.5mm，气溶胶颗粒的速度约为400m/s时，两测量值之间的持续时间约为6.25μs。为了在上述点的测量值之间获得所需的无歧义性，根据本发明的第一方面，第一激光装置是一种双色连续波激光装置，以266至532nm的波长工作。照射到第一点的光的波长为532nm，而照射到第二点的光的波长为266nm。后一种波长也是使生物气溶胶颗粒显示荧光。
例如，第二激光装置是准分子激光装置，工作在308nm的波长下，且在正确触发之后，确保仅生物气溶胶颗粒电离。为此，如前所述，该激光装置的脉冲由荧光检测器触发。UV准分子激光的使用意味着为了使MALDI基础上的所述方法的性能成为可能，气溶胶颗粒必须具有基质(matrix)。根据本发明的另一方面，在吸入ATOFMS过程中或就在此之前，气溶胶颗粒通过蒸发/冷凝或升华/冷凝而具有这种基质。如果使用IR(红外)激光装置，这不是必需的。
附图说明
现在将在参照附图的示例性实施例基础上更详细地解释本发明的方法和装置，其中：
图1示出了本发明的可以使用用于检测和识别生物气溶胶颗粒的方法的装置；
图2示意性地示出了ATOFMS中颗粒流穿过的一部分距离，其中示出了颗粒受到相应激光辐射的位置。
具体实施方式
图1中所示的ATOFMS以剖面形式示出，即垂直于所述气溶胶颗粒流。所述气溶胶颗粒流基本上沿线2而行。气溶胶颗粒从空气中抽入到ATOFMS中，且在该装置的入口部分在高温下设有源于容易升华的物质的涂层。为此，众所周知例如使用吡啶羧酸或芥子酸。然后通过气动透镜系统、收缩喷嘴和撇离器使气溶胶颗粒成束。图2图示出气溶胶颗粒流是如何从上述入口部分的喷嘴3经300μm的出口4以及分别为400μm和300μm的撇离器开口5和6进入ATOFMS的中央空间7。这一空间被气溶胶颗粒一个接一个地以特定的相互距离连续穿过。在本示例中，气溶胶颗粒进入ATOFMS的中央空间的速度约为400m/s。需穿过的相应空间中的压力分别为1、10-3和10-6毫巴。在所述中央空间7中，气溶胶颗粒流首先受到第一激光装置8的照射。在图1中，这是DPSS(二极管泵激式固态)激光器，它是双色连续波激光器这一特定类型。这种激光装置8将波长为532nm的光束发送到空间7，即在点9处，以及将波长为266nm的光束发送到点10的位置处(见图2)。两种波长的光被分色镜11和12偏转。点9沿线2的方向的宽度约为50μm，且在与线2垂直的平面内该点的直径约为150μm。气溶胶颗粒暴露的时间约为125ns。点10沿线2的方向的宽度约为500μm，在与线2相垂直的平面内该点的直径约为150μm。在点9上暴露的气溶胶颗粒使所述光散射，而在点10上气溶胶颗粒使所述光散射并发射荧光辐射，只要气溶胶颗粒是生物起源。气溶胶颗粒散射的光线由检测器13和14检测。为此，这些检测器设有光电倍增管。检测器13直接检测从点9散射的光线。经半透反射镜15，检测器14检测从点10散射的光线，而荧光辐射由该反射镜传递到荧光检测器16。因为反射镜15总是传递一小部分266nm的散射辐射，所以荧光检测器16包含在入口的通带滤光器17，该滤光器将这一波长进一步过滤掉，但传递在300至500nm范围内的荧光辐射。点10沿线2的方向的宽度较大。在第一个125ns内，相当于点9的宽度，在检测器14中检测气溶胶颗粒的散射光，所以在连续观测点9和10的散射光的基础上，可以确定相应的气溶胶颗粒的尺寸。在下一125ns内，所述荧光检测器16被检查相应的气溶胶颗粒实际上是否是生物起源的检测器所释放。气溶胶颗粒在点10上被辐射的过程中的剩余时间提供了提高荧光测量的可靠性的时机。除了各反射镜18、19和20之外，荧光检测器16还包含衍射光栅21和CCD录像机22。当记录荧光辐射时，控制信号通过摄像控制单元23送往触发电路24。而且来自两检测器13和14的输出信号也送往该触发电路24。在触发电路24中，确定气溶胶颗粒是否是具有特定大小的生物起源，此后，触发信号送往受激准分子激光器25。换言之，特定尺寸的气溶胶颗粒一被认为是生物起源，就触发受激准分子激光器装置。即，受激准分子激光器装置提供激光脉冲，该脉冲影响点26上的相应生物气溶胶颗粒的电离。为了确保所述生物气溶胶颗粒实际上电离，该点相对较大：在线2的方向上约300μm，在与线2相垂直的平面内约500μm。点10和26之间的距离调节为所述气溶胶颗粒的速度和产生触发信号所需的时间。经上述分色镜11、12，所述准分子激光脉冲传递到ATOFMS中的中央空间7中。这样，电离的生物气溶胶颗粒通过电极27以普通的方式加速并且偏转，且经透镜28和偏转装置29到达离子检测器30。在显示器31上示出产生的质谱结果。
本发明不限于参照附图所示的示例性实施例，而包含其所有的改进，当然，只要它们落在所附权利要求的保护范围内。因此，可以使用其他类型的激光装置，且当作为用于识别生物起源的气溶胶颗粒的辅助手段的尺寸确定不再认为必须时，则不必使用双色激光装置。而且，代替具有衍射光栅和CCD录像机的荧光检测器16，可以使用任何其他公知的类型；衍射光栅和CCD录像机可以例如由具有通带滤光器的“选通PMT”(gated PMT)代替。所述点的尺寸仅通过示例给出；它们当然可以不同地选择，正如说明书中提及的其他参数。因为光线被气溶胶散射或发射到各方，所以可以设置数个检测器。严格而言，不是绝对必须使用检测器13和14。荧光检测器足以用于识别生物起源的气溶胶颗粒，并触发受激准分子激光器。然而，在实践中，这种实施例的可靠性是不够的。
Claims (7)
1.一种用于检测和识别空气中的生物气溶胶颗粒的方法，其中在ATOFMS(气溶胶微粒飞行时间质谱分析仪)中，通过荧光技术选择颗粒流中的生物气溶胶颗粒，且仅使从其中检测出荧光辐射的所选生物气溶胶颗粒被电离，此后接着检测生成的离子，并识别出生物气溶胶颗粒。
2.如权利要求1所述的方法，其特征在于，生物气溶胶颗粒的选择是通过激光辐射进行的，所述激光辐射由第一激光装置产生，其波长在生物气溶胶颗粒的特定物质中产生荧光，此后，通过荧光检测器选择生物气溶胶颗粒，然后触发第二激光装置发射光束，该光束的波长使仅由荧光检测器选择的生物气溶胶颗粒电离。
3.如权利要求1或2所述的方法，其特征在于，在吸入ATOFMS过程中或就在此之前，气溶胶颗粒通过蒸发/冷凝或升华/冷凝而具有基质，此后，所选择的生物气溶胶颗粒在MALDI(基质辅助的激光解吸/电离)基础上电离，此后接着检测离子并识别出生物气溶胶颗粒。
4.一种利用ATOFMS的装置，其特征在于，设置有用于发射光束的第一激光装置，该光束的波长在生物气溶胶颗粒的特定物质中产生荧光，该装置还具有用于检测荧光辐射的相应荧光检测器，以及由所述检测器触发、用于发射光束的第二激光装置，该光束的波长使由荧光检测器选择的生物气溶胶颗粒电离。
5.如权利要求4所述的装置，其特征在于，所述第一激光装置是一种连续波激光装置，所述第二激光装置是一种脉冲激光装置。
6.如权利要求4或5所述的装置，其特征在于，所述第一激光装置是一种双色连续波激光装置，在266和532nm的波长下运行。
7.如权利要求4或5所述的装置，其特征在于，所述第二激光装置是一种准分子激光装置，在308nm的波长下运行。
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| JP4145651B2 (ja) | 2008-09-03 |
| IL156444A (en) | 2009-07-20 |
| AU2002226802A1 (en) | 2002-07-08 |
| WO2002052246A2 (en) | 2002-07-04 |
| US6806464B2 (en) | 2004-10-19 |
| EP1342256A2 (en) | 2003-09-10 |
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