CN115667890B - 基于空间梯度的荧光计 - Google Patents
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Abstract
一种基于空间梯度的荧光计,其特征在于信号处理器或处理模块,该信号处理器或处理模块被配置为:接收信令,该信令包含关于从液体中的荧光团反射并且由线性传感器阵列感测的光的信息,该线性传感器阵列具有长度以及多行和多列的光学元件;以及基于所接收的信令确定包含关于液体的荧光团浓度的信息的对应信令,液体的荧光团浓度取决于沿线性传感器阵列的长度所反射和感测的光的空间梯度。
Description
相关申请的交叉引用
本申请要求于2020年5月20日提交的临时专利申请序列号63/027,587(911-023.9-1-1/N-YSI-0045US01)、于2020年5月21日提交的临时专利申请序列号63/028,013(911-023.010-1-1/N-YSI-0046US02)以及于2020年5月22日提交的临时专利申请序列号63/028,723(911-023.011-1-1/N-YSI-0047US02)的权益,其全部通过整体引用并入本文。
技术领域
本发明涉及一种用于测量液体中的感兴趣物种的浓度的荧光计;并且更具体地涉及一种用于使用基于非强度(即,幅度)的测量来测量液体中的荧光团的浓度的荧光计。
背景技术
光学荧光现象通常用于环境水质监测,因为这种技术可以实现为紧凑的现场坚固的传感器。基于荧光的感测包括激发光源(在指定光学波长下),激发光源用于光学激发感兴趣水参数并且重新发射特定于感兴趣水参数的光(在较长光学波长下)。
已知荧光计通过测量返回荧光信号的幅度来测量水物种的浓度。基于幅度的测量存在多个问题:
1)激励源的劣化
典型的激励源包括LED、激光二极管或灯,所有这些都在使用过程中受到强度下降的影响。用于处理源劣化的选项是有限的。一个选项是包括参考检测器,参考检测器将消除/取消劣化的影响,但增加了传感器电路的复杂性,并且需要附加的光机空间。第二选项是定期重新校准传感器,这必然会限制现场部署的持续时间。
2)激励源的热漂移
上述所有光源对温度具有不可忽略的响应,即,光输出功率随环境温度变化而变化。这再次依赖于参考检测器或某种复杂的电气或嵌入式软件补偿方案,在传感器性能方面带来了真正的问题。此外,温度补偿需要温度传感器的某种测量,其通常由板载(即,位于电路内部)温度传感器实现,这需要附加的电路系统和物理空间。
3)干扰物种
基于荧光的传感器可能遭受光学干扰,其中存在其他竞争物种,其他竞争物种可能在相同的相应目标激发和/或发射波长下吸收,导致荧光幅度的降低。
4)光学机械配置
传统的荧光感测技术由于荧光信号的不良/低效捕获而导致灵敏度不良(尤其是现场可部署传感器)。现有荧光传感器通常采用单个激发光源和利用了光敏元件的单个(点状)发射接收器。无论使用何种特定光敏元件或激发光源,已知的现有技术都没有针对荧光的高效捕获而进行光学机械配置,从而导致检测极限受损。
5)内滤(Inner Effect)效应(IFE)——范围限制效应
已知的现有技术存在以下问题:在低浓度下,荧光信号与物种浓度近似成比例。然而,随着浓度的增加,信号达到最大值,随后信号随着浓度的升高而降低。在这点上,传统的荧光计是模糊的双值,这表示,对于任何特定测量荧光信号,有两种可能的浓度,一种是高,一种是低。对于这些已知的荧光计,无法区分这两种可能的结果。
已知文献
已知文献涉及使用2-D阵列来估计浓度梯度的荧光,并且主要发现的简要总结如下:
已知的现有技术公开了用于“在空间和时间中确定浓度梯度”的2-D阵列。这里,(“扩散驱动”)浓度梯度由荧光信号的局部分布和幅度确定。这是一种基于幅度的技术,因为由任何特定阵列元件报告的信号仅与“局部”荧光的量(即,特定单个阵列元件处的局部信号)成比例,其中荧光的局部幅度被理解为与局部荧光团浓度密度成比例。
此外,参见2008年4月7日提交的PCT/US2008/059575(其公开了一种用于高通量浊度测量的系统和方法)以及A Singh等人的题为“The performance of 2D arraydetectors for light sheet based fluorescence correlation spectroscopy”的文章。
发明内容
本发明明显不同于上述现有技术:
例如,本发明使用空间梯度(比尔定律的结果)来确定单个固定/准静态浓度,其中浓度随时间的变化被理解为比信号获取所需要的时间慢得多。换言之,根据本发明的传感器的空间梯度是比尔定律的结果,而不是荧光团浓度的某种改变/变化的空间分布的结果。
此外,本发明避免了与基于幅度的荧光测量相关联的很多问题,同时提供了能够大大增强信号捕获和IFE消除的光学机械配置。本发明采用线性光电二极管阵列(然而,本发明不限于光电二极管技术,例如,也可以使用线性CCD或CMOS阵列)。线性阵列允许基于非强度的荧光确定。这些测量是空间相关的,其主要思想是,光信号将按照比尔定律跨线性阵列进行衰减,从而产生“空间梯度”。该空间梯度包含关于荧光物质的浓度的信息。
本发明的关键要素具体涉及使用线性传感器阵列来评估沿线性传感器阵列的长度的信号的空间梯度。信号的空间梯度提供荧光团浓度的评估,与已知的基于幅度的方法相比,这提供很多优点,包括:
-抗源劣化/漂移,
-无校准感测,
-抗荧光带干扰,
-增强的信号灵敏度,以及
-IFE校正。
其他实现
上述“空间梯度”方法要求阵列中的每个光学元件都个体可寻址。然而,存在一种可能的设计变型,其涉及添加透射光电二极管(位于阵列的端部处,与源相对)并且以电并联配置连接所有线性阵列元件。该设计变型将进一步提高低信号灵敏度,从而进一步提高最小检测极限,同时保持传感器执行漂移和IFE校正的能力。
最后,除了基于幅度的方法之外,另一变型可以包括空间梯度方法,以提供补充信息。这里,基于梯度的方法可以用于标识激励劣化,而基于幅度的方法可以被用于支持低信号检测。具体实施例
根据一些实施例,本发明可以包括一种装置或采取一种装置的形式,该装置的特征在于信号处理器或处理模块,该信号处理器或处理器模块被配置为:
接收信令,该信令包含关于从液体中的荧光团反射并且由线性传感器阵列感测的光的信息,该线性感测阵列具有长度以及多行和多列的光学元件;以及
基于所接收的信令确定包含关于液体的荧光团浓度的信息的对应信令,该荧光团浓度取决于沿线性传感器阵列的长度所反射和感测的光的空间梯度。
该装置可以包括以下附加特征中的一个或多个:
该装置可以包括线性传感器阵列。
线性传感器阵列可以包括线性光电二极管阵列、线性CCD阵列或线性CMOS阵列、以及具有三维圆柱形阵列的多行和多列的光学元件的闭式柱状传感器阵列。
空间梯度可以由线性阵列算法确定,该线性阵列算法定义荧光团浓度[c]、沿线性传感器阵列的长度或位置(l)、物种吸收系数(α)和沿线性传感器阵列的阵列光学元件的信号(S(I))之间的关系。
线性阵列算法采用以下等式的形式:
y=mx+b,
其中
y=-ln(S(I)),
mx=α[c]I,并且
b=-ln([c]AT0)。
线性阵列算法基于比尔定律。
该装置可以包括基于空间梯度的荧光计或采取基于空间梯度的荧光计的形式。
该装置可以包括准准直光源,该准准直光源具有对应长度并且被配置为沿线性传感器阵列的长度提供包括准准直光的光。
信号处理器或处理模块可以被配置为:基于跨线性传感器阵列、包括沿线性传感器阵列的长度和/或宽度而感测的光信号的衰减来确定荧光团浓度。
线性传感器阵列可以包括个体可寻址的光学元件的二维阵列。
多行或多列的光学元件可以并联连接并且由信号处理器或处理模块可寻址;该装置可以包括位于线性传感器阵列的与光源相对的端部处的透射光电二极管,透射光电二极管被配置为响应于从荧光团反射的光并且提供包含关于光的信息的透射光电二极管信令;并且信号处理器或处理模块可以被配置为接收光电二极管信令并且针对漂移或内滤效应(IFE)校正对应信令。
基于空间梯度的荧光计
又例如,并且根据一些实施例,本发明可以包括基于空间梯度的荧光计或采取基于空间梯度的荧光计的形式,该基于空间梯度的荧光计的特征在于准准直光源、线性传感器阵列和信号处理器或处理模块。
准准直光源具有长度,并且可以被配置为向液体样品提供准准直光。
线性传感器阵列具有对应长度以及多行和多列的光学元件,并且可以被配置为沿准直光源的长度感测从液体样品中的荧光团反射的光并且提供包含关于从荧光团反射的光的信息的信令。
信号处理器或处理模块可以被配置为:
接收信令;以及
基于所接收的信令确定包含关于液体的荧光团浓度的信息的对应信令,该荧光团浓度取决于沿线性传感器阵列的对应长度所反射和感测的光的空间梯度。
基于空间梯度的荧光计还可以包括上述特征中的一个或多个。
方法
根据一些实施例,本发明可以包括一种方法,该方法的特征在于:
利用信号处理器或处理模块接收信令,该信令包含关于从液体中的荧光团反射并且由线性传感器阵列感测的光的信息,线性传感器阵列具有长度以及多行和多列的光学元件;以及
利用信号处理器或处理模块基于所接收的信令确定包含关于液体的荧光团浓度的信息的对应信令,该荧光团浓度取决于沿线性传感器阵列的长度所反射和感测的光的空间梯度。
该方法还可以包括上述特征中的一个或多个。
计算机可读存储介质
根据本发明的一些实施例,本发明还可以采取计算机可读存储介质的形式,计算机可读存储介质具有用于执行上述方法的步骤的计算机可执行组件。计算机可读存储介质也可以包括上述特征中的一个或多个。
优势
本发明提供了优于现有技术中的当前已知技术的明显优点,如下:
1)本发明通过空间梯度(跨线性阵列检测器的长度而变化的荧光信号,符合比尔定律)(参见图1)而不是通过荧光信号的幅度(浓度确定算法(参见图5))来确定荧光团浓度。因此,它不受光源强度中的适度变化的影响。这表示,空间梯度不受源劣化、源热响应或源驱动条件(诸如LED驱动电流)的变化的影响。然而,必须存在不可忽略的信号,即,必须有一些可测量量的光入射到阵列上以形成空间梯度。此外,这些元素需要个体可寻址以解析空间信息。本发明不限于任何特定线性阵列检测器技术;可以使用线性光电二极管、CCD或CMOS阵列。
2)本发明不受源劣化/漂移的影响,能够进行无校准部署,从而延长每次部署的长度。
3)线性传感器阵列提供了大得多的总有效面积以捕获返回荧光。更重要的是,沿光轴在最重要的维度上的有效面积更大(通常使用准准直激励源,其主要沿通常称为“光轴”的单个轴发射辐射)(参见图1)。荧光捕获的增加大大增强了信号灵敏度,这进而显著提高了荧光物种检测的最小极限。
4)正如基于梯度的方法不受激励功率中的适度变化的影响一样,它也不受某些类型的干扰的影响。吸收荧光信号但不吸收激励信号的任何干扰物质(荧光带干扰)将不会影响信号梯度,并且因此不会妨碍荧光团浓度的任何评估。注意,空间梯度方法不能解决吸收激励信号的任何干扰物种(激励带干扰),因为这会影响信号梯度的特征。
5)虽然传统荧光计的荧光幅度具有模糊的双值响应(由于IFE),但空间梯度方法的情况并非如此,空间梯度方法的响应随着浓度的增加而是单调的(参见图4)。空间梯度方法实现了实时内滤效应(IFE)校正。[对于已知的现有技术,内滤效应校正的常用方法涉及在现场部署之后通过实验室分析进行后处理]。IFE校正大大增强了高浓度感测范围(参见图3)。
附图说明
附图(不一定按比例绘制)包括图1-图8,如下所示:
图1是遵循比尔定律的荧光“空间梯度”的侧视图(在TraceProTM中模拟)。
图2包括图2A和图2B,示出了荧光梯度的空间绘图和强度图(在TraceProTM中模拟)。
图3是具有和没有IFE校正的传感器响应与相对浓度的关系图[示出了10倍增强的检测范围](在TraceProTM中模拟)。
图4是具有和没有IFE校正[消除双值问题]的传感器响应与相对浓度的关系图(在TraceProTM中模拟)。
图5是根据本发明的一些实施例的根据空间梯度确定浓度的算法。
图6是根据本发明的一些实施例的装置的框图,该装置包括基于空间梯度的荧光计。
图7是根据本发明的一些实施例的具有长度以及多行和多列的光学元件的线性传感器阵列的框图。
图8是根据本发明的一些实施例的准准直光源的三维透视图,该准准直光源相对于线性传感器阵列提供准准直光。
为了减少图中的混乱,图中的每个图不一定包括其中示出的每个元素的每个附图标记。
具体实施方式
图6示出了根据本发明的包括基于空间梯度的荧光计的装置10,装置10具有准准直光源20、线性传感器阵列30和信号处理器或处理模块40。
信号处理器或处理模块40可以被配置为
接收信令,该信令包含关于从液体中的荧光团反射并且由线性传感器阵列30感测的光Lr(图8)的信息,线性传感器阵列30具有长度L以及例如,如图7所示的多行和多列的光学元件……(r1,c1;r1,c2;r1,c3;r1,c4;r1,c5;r1,c6;r1,c7;r1,c8;……;r1,cn;r2,c1;r2,c2;r2,c3;r2,c4;r2,c5;r2,c6;r2,c7;r2,c8;……;r2,cn;r3,c1;r3,c2;r3,c3;r3,c4;r3,c5;r3,c6;r3,c7;r3,c8;……;r3,cn;……;rn,c1;rn,c2;rn,c3;rn,c4;rn,c5;rn,c6;rn,c7;rn,c8;……;rn,cn)。
基于所接收的信令确定包含关于液体的荧光团浓度的信息的对应信令,该荧光团浓度取决于沿线性传感器阵列30的长度L所反射和感测的光的空间梯度。
线性传感器阵列30
例如,装置10可以包括线性传感器阵列30,例如,诸如线性光电二极管阵列、线性电荷耦合器件(CCD)阵列、线性CMOS阵列。又例如,线性传感器阵列30可以包括二维阵列的多行和多列的光学元的,例如,如图7所示,光学元件是个体可寻址的。线性传感器阵列在本领域中是已知的,并且本发明的范围不旨在局限于现在已知或将来开发的任何特定类型或种类。
例如,线性传感器阵列在以下美国专利No.9,020,202、8,022,349、7,956,341、7,040,538、5,252,818和4,193,057中公开,其全部通过引用并入本文。
光源20
例如,装置10可以包括光源20,光源20被配置为提供包括准准直光的光Lc(图8),光Lc沿线性传感器阵列30的长度通过与光源20和线性传感器阵列30相关布置的液体样品L,以便将光Lr从被监测或测试的液体样品中的荧光团反射到线性传感器阵列30上。参见图8。例如,光可以被径向和向后反射,即,反向散射的反射光或辐射。
如本领域技术人员将理解的,准准直光源在本领域中是已知的,并且本发明的范围不旨在局限于现在已知或将来开发的任何特定类型或种类的光源。
信号处理器或处理模块40
例如,信号处理器或处理模块40可以被配置为基于跨线性传感器阵列而感测的光信号的空间梯度来确定荧光团浓度,例如,与图5中所述的一致。
在备选实施例中,多行或多列的光学元件可以并联连接并且由信号处理器或处理模块40可寻址;装置10可以包括位于线性传感器阵列30的与光源20相对的端部处的透射光电二极管30a,透射光电二极管30a被配置为响应于从荧光团反射的光并且提供包含关于该光的信息的透射光电二极管信令;并且信号处理器或处理模块40可以被配置为接收光电二极管信令并且针对漂移或内滤效应校正对应信令。
信号处理功能的实现
例如,信号处理器或处理模块40的功能可以使用硬件、软件、固件或其组合来实现。在典型的软件实现中,信号处理器40将包括一个或多个基于微处理器的架构,该架构具有例如至少一个信号处理器或微处理器。本领域技术人员将能够用合适的程序代码编程这样的基于微控制器或基于微处理器的实现以执行本文中公开的信号处理功能,而无需过度实验。
本发明的范围不限于使用现在已知或将来开发的技术的任何特定实现。本发明的范围旨在包括将(多个)信号处理器的功能实现为独立处理器、信号处理器或信号处理器模块、以及单独的处理器或处理器模块、以及其某种组合。
例如,装置10还可以包括例如其他信号处理器电路或组件,总体上用50表示,包括随机存取存储器或存储器模块(RAM)和/或只读存储器(ROM)、输入/输出设备和控制器、连接它们的数据和地址总线、和/或至少一个输入处理器和至少一个输出处理器,例如,这将为本领域技术人员所理解。
又例如,信号处理器40可以包括信号处理器和包含计算机程序代码的至少一个存储器的某种组合,或者采取这种组合的形式,其中信号处理器和至少一个存储器被配置为引起系统实现本发明的功能,例如,响应于所接收的信令并且基于所接收的信令确定对应信令。
内滤效应(IFE)
如本领域技术人员将理解的,IFE是一种荧光光谱现象,例如,其中由于靠近入射光束的荧光团对激发光的吸收,在浓溶液中看到的荧光发射减少,这显著地减少了到达远离入射光束的样品的光。
如本领域技术人员将理解的,用于校正IFE的技术在本领域中是已知的,并且本发明的范围不旨在局限于现在已知或将来开发的任何特定类型或种类。
比尔定律
如本领域技术人员所理解的,比尔定律由以下关系式定义:
A=εbC,
其中
A=吸光度,
ε=摩尔吸收率,
b=光路长度,以及
C=浓度,
荧光团
如本领域技术人员将理解的,荧光团是一种荧光化合物,其在激发时可以重新发光。荧光团通常含有若干组合的芳香基团、或具有π键的平面或环状分子。
例如,荧光团有时用作流体中的示踪剂,用作对某些结构染色的染料,用作酶的底物,或用作探针或指示剂(当荧光受到环境因素(诸如极性或离子)的影响时)。
本发明的范围不旨在局限于现在已知或将来开发的任何特定类型或种类的荧光团。
应用
本发明具有应用,例如在淡水应用的水质监测的基本参数中以及在饮用水监测中。
本发明的范围
虽然已经参考示例性实施例描述了本发明,但是本领域技术人员将理解,在不脱离本发明的范围的情况下,可以进行各种改变,并且可以用等效物代替其元素。此外,在不脱离本发明的基本范围的情况下,可以进行修改以使特定情况或材料适应本发明的教导。因此,意图在于,本发明不限于本文中公开的作为实施本发明的最佳模式的(多个)特定实施例。
Claims (18)
1.一种装置,包括:
信号处理器或处理模块,被配置为:
接收信令,所述信令包含关于从液体中的荧光团反射并且由线性传感器阵列感测到的光的信息,所述线性传感器阵列具有长度以及多行和多列的光学元件;以及
基于所接收的信令,确定包含关于所述液体的荧光团浓度的信息的对应信令,所述荧光团浓度取决于沿所述线性传感器阵列的所述长度所反射并感测到的所述光的空间梯度,
其中所述线性传感器阵列中的每个光学元件是个体可寻址的。
2.根据权利要求1所述的装置,其中所述信号处理器或处理模块被配置为:基于跨所述线性传感器阵列的长度而感测到的光信号的衰减来确定荧光团浓度。
3.根据权利要求1所述的装置,其中所述线性传感器阵列包括线性光电二极管阵列、线性CCD阵列或线性CMOS阵列。
4.根据权利要求1所述的装置,其中所述线性传感器阵列包括闭式柱状传感器阵列,所述闭式柱状传感器阵列具有三维圆柱形阵列的所述多行和所述多列的所述光学元件。
5.根据权利要求1所述的装置,其中所述空间梯度由线性阵列算法确定,所述线性阵列算法定义所述荧光团浓度[c]、沿所述线性传感器阵列的所述长度或位置(l)、物种吸收系数(α)以及沿所述线性传感器阵列的阵列光学元件的信号(S(I))之间的关系。
6.根据权利要求5所述的装置,其中所述线性阵列算法基于比尔定律。
7.根据权利要求1所述的装置,其中所述装置是基于空间梯度的荧光计。
8.根据权利要求1所述的装置,其中所述装置包括准准直光源,所述准准直光源具有对应长度并且被配置为沿所述线性传感器阵列的所述长度提供包括准准直光的所述光。
9.根据权利要求1所述的装置,其中所述信号处理器或处理模块被配置为:基于跨所述线性传感器阵列、包括沿所述线性传感器阵列的所述长度和宽度而感测到的光信号的衰减,确定所述荧光团浓度。
10.根据权利要求1所述的装置,其中所述光学元件由所述信号处理器或处理模块个体可寻址。
11.根据权利要求10所述的装置,其中:
所述多行或所述多列的所述光学元件并联连接并且由所述信号处理器或处理模块可寻址;
所述装置包括透射光电二极管,所述透射光电二极管位于所述线性传感器阵列的与所述光源相对的端部处,所述透射光电二极管被配置为响应于从所述荧光团反射的所述光并且提供包含关于所述光的信息的透射光电二极管信令;以及
所述信号处理器或处理模块被配置为接收所述光电二极管信令并且针对漂移或内滤效应校正对应信令。
12.一种方法,包括:
利用信号处理器或处理模块接收信令,所述信令包含关于从液体中的荧光团反射并且由线性传感器阵列感测到的光的信息,所述线性传感器阵列具有长度以及多行和多列的光学元件;以及
基于所接收的信令,利用所述信号处理器或处理模块确定包含关于所述液体的荧光团浓度的信息的对应信令,所述荧光团浓度取决于沿所述线性传感器阵列的所述长度所反射并感测到的所述光的空间梯度,
其中所述线性传感器阵列中的每个光学元件是个体可寻址的。
13.根据权利要求12所述的方法,其中所述方法包括:将所述线性传感器阵列配置为线性光电二极管阵列、线性CCD阵列或线性CMOS阵列。
14.根据权利要求12所述的方法,其中所述方法包括:基于跨所述线性传感器阵列的所述长度而感测到的光信号的衰减,确定所述荧光团浓度。
15.根据权利要求12所述的方法,其中所述方法包括:配置光源以提供所述光,该配置步骤包括使用准准直光源来提供准准直光。
16.一种基于空间梯度的荧光计,包括:
准准直光源,具有长度并且被配置为向液体样品提供准准直光;
线性传感器阵列,具有对应长度以及多行和多列的光学元件,并且被配置为:沿所述准直光源的所述长度感测从所述液体样品中的荧光团反射的光,并且提供包含关于从所述荧光团反射的所述光的信息的信令;以及
信号处理器或处理模块,被配置为:
接收所述信令;以及
基于所接收的信令,确定包含关于所述液体的荧光团浓度的信息的对应信令,所述荧光团浓度取决于沿所述线性传感器阵列的对应长度所反射并感测到的所述光的空间梯度,
其中所述线性传感器阵列中的每个光学元件是个体可寻址的。
17.根据权利要求16所述的基于空间梯度的荧光计,其中所述线性传感器阵列包括线性光电二极管阵列、线性CCD阵列或线性CMOS阵列。
18.根据权利要求16所述的基于空间梯度的荧光计,其中所述空间梯度由线性阵列算法确定,所述线性阵列算法定义所述荧光团浓度[c]、沿所述线性传感器阵列的所述长度或位置(I)、物质吸收系数(α)与沿所述线性传感器阵列的阵列光学元件的信号(S(I))之间的关系。
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