基于定量离心分层技术的快速血液检测关键技术与装置研究
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摘要
目的：血液中各类血细胞含量是反映人体健康程度的重要参数。血液流经人体各个组织器官，其成分变化对于人体机能和组织功能的实现产生重大影响，而机体的病变也会影响血液各组分，因此，对人体血细胞进行分析具有重大意义。目前临床检验设备主要是基于湿化学方法，其设备普遍存在检验速度慢、体积大、需大量液体试剂的配合，现场应用受限等问题。本文研究目的是研究一种基于定量离心分层技术（干式化学法）的快速血液检测关键技术及装置，实现定量检测常规血细胞参数，使其适合现场条件、急诊情况或基层医疗单位使用。
方法：首先，在研究总结了前人的研究成果基础上提出血细胞分析检测方法：离心、检测相分离的基于血液棕黄色层定量离心分层技术（QBC）的方法。为离心机设计可快速拆卸的专用离心盘，用来配合干式血液检测毛细管的离心，而在更换离心盘后，该离心机可用于常规的临床检验实验室用途，这样可以做到一机多用。该算法对传统检测方法进行了改进：首先，采用彩色线阵CCD结合自行设计的成像透镜对待测毛细管实现一次成像，大大提高检测精度与速度；其次，在用蓝光激发荧光照射时，使毛细管沿其轴向转动10个位置检测取平均值，减小了由于离心过程中各个界面分层不均带来的影响；最后，通过医学实验对系统的可靠性和可行性进行了检验分析。
内容：本论文主要包括以下几个方面的研究工作：
（1）进行了光路及CCD图像采集模块的研究。在光路及CCD图像采集模块中，采用LED冷光源增强了环境适应性；应用彩色线阵CCD不需额外的滤光片，在信号处理电路中采用相关双采样法及视频信号处理专用芯片AD9826，简化了系统的结构；应用CPLD可编程逻辑器件实现CCD驱动的时序，简化程序，提高应用性。
（2）对图像采集后的信号处理算法进行了研究。该部分主要是确定各层分层的确切位置，在红光源照射下，确定全血体积、标定点及浮子位置；在蓝光源照射下，重点区分中间三层（白细胞和血小板层）的信息。中间三层的边界确定采用红色图和绿色图相互比照的方法。确定了算法规则及血常规参数的转化。
（3）对系统毛细管进排样、检测控制模块进行了研究。设计了毛细管进排样的机械结构，中央处理单元采用DSP数字信号处理器，减少外围电路，进行了进排样精度及检测精度的实验，系统稳定性良好。
（4）在研究的新型检测方法基础上，结合光路技术和电路技术等，构建了“基于定量离心分层技术的快速血液检测系统”。本文详细叙述了系统三个组成部分：光路检测结构、毛细管进排样结构及数据处理算法的实现。
（5）设计并开展了一项医学实验，对系统的可靠性和可行性进行检验。在与QBC Autoread plus和Sysmex XT-1800i两类仪器的对比中，以红细胞压积（HCT）、血红蛋白（HGB）、粒细胞（GRAN）、淋巴细胞及单核细胞（LM）、白细胞（WBC）和血小板（PLT）为对比参数，相关系数分别为0.99、0.98、0.95、0.93、0.96、0.94和0.99、0.97、0.986、0.95、0.975、0.94，达到了令人满意的效果。
结果：
提出了基于干式化学法的快速血细胞检测方法，并在以数字信号处理器（DSP）为核心控制的系统中，结合光路检测技术、电路控制技术和数据处理技术，构建了“基于定量离心分层技术的快速血液检测系统”检验平台；制作完成实验样机一套，通过对大量样本的实验测试，验证了系统的可靠性和可行性。针对基于干式化学法的血液检测系统的毛细管进排样系统，申请发明专利一项。
结论：
本系统完全突破了传统的血液检测概念，实现了基于干式化学法的检测，相比湿式检测具有灵活、不需要液体试剂处理过程。本课题实现了现场条件下仪器携行性、抗震性好、检测结果快速准确的功能。一方面及时准确的检测结果为伤病员的抢救赢得了时间；另一方面，系统结构紧凑、运营成本低，同样适合基层医疗单位使用。“基于定量离心分层技术的快速血液检测系统”性能优良、使用方便，运用该系统可实现血细胞HCT、HGB、RBC、PLT、MCHC、GRAN、%GRAN、LM和%LM9个参数的定量测量，并具有自校准和报警提示、输出存储与打印等功能。在现场血液快速检测和基层急诊中有较广泛的应用价值，特别是在战时状态进行常规血细胞检测具有不可替代的价值。
研究意义：
1）样本无需制备，可迅速展收，适合现场快速检验；
2）检测速度快，除去离心时间（5分钟），进样、检测时间≯60s，操作简便，无需复杂维护，携行性好；
3）结构紧凑，体积小，环境适应性强，使用综合成本低。
本研究特色及创新之处：
1）离心、检测分离，除去离心的5分钟，每个样本检测时间在60秒内，大大提高检测速度，尤其提高多样本的检测速度（以20样本为例，本系统与QBCAutoread plus和QBC star相比，三者检测时间分别是25分钟、45分钟和120分钟）；
2）双光源检测、毛细管轴向转动，高分辨率的彩色线阵CCD图像传感器一次成像，提高检测精度；
3）冷光源LED功耗低、控制简单、寿命长，工作温度范围广，环境适应性、抗震动和抗干扰能力强；
4）采用DSP作为核心控制器，实时、快速地实现各种数字信号的处理，软件方面把对两个电机的控制以任务的形式进行调度，提高了系统的反应速度；
5）检测模块功能独立，结构大大简化，体积小，重量轻，携行性好，有效降低成本，提高可集成性，优于国内外的同类产品。
Objective: The contents of various blood cells in the blood are important parametersto reflect the health of human body. The blood flows through all the tissues andorgans and any change of its contents will exercise great influence on the realizationof the functions of human bodies and tissues. Likewise, any pathological change inthe functions will also influence blood contents. So analysis of the blood cells inhuman bodies is of great importance. Currently, clinical detection equipments, whichare mainly based on wet-type chemical method, have some disadvantages in commonsuch as low speed of detection, big size, requiring large amount of liquid reagents togo with, restriction in field application, and so on. This paper aimed to research thekey technology and equipments of rapid blood detection based on QBC technique, adry-type chemical method, so as to achieve the quantitative detection of normal cellparameters and find its application in field occasion, emergency situation andbasic-level medical units.
Methods: Based on the research and summary of achievements previously made, anew method to detect blood cells was proposed as QBC technique which realized theseparation of centrifugation and detection on the basis of quantitative buffy coat. Acentrifugal disc, which could be disassembled very conveniently and quickly, wasdesigned specifically for the centrifuge, which could also be used to routine clinicallaboratory when the special centrifugal disc was replaced so as to realize more thanone purpose in a machine. The method has made some improvements on thetraditional test methods as follows: firstly, color linear array CCD, combined withself-designed imaging lens was adopted to fulfill instant photograph of theto-be-tested capillaries, which greatly improved the accuracy and speed of detection;secondly, the capillaries were designed to rotate along the axial direction when theywere irradiated with fluorescence excited by blue lights, then they were detected at10times to get their average value, as reduced the effect brought about by unevenness ofthe layering of interfaces in the course of centrifugation; finally the system was examined to prove its feasibility and reliability through medical experiments.
Contents: This paper is mainly composed of the researches in the following aspects:
1) Research on optical path and CCD image collection module. In the module ofoptical path and image collection, LED cold light source was employed to enhance itsenvironmental suitability; the configuration of the system was simplified by usingcolor linear array CCD to replace additional spectral filter, and by adopting coupledouble sample (CDS) method and AD9826, a special-purpose video-signal processingchip in the signal processing electric circuit; the application of CPLD, aprogrammable logic device, was the time sequence to realize the drive of CCD, whichcould simplify procedures and improve the applicability.
2) Research on the signal processing algorithm after the collection of images.This part was intended to determine the definite position of each layer, and in theirradiation of red light source, to decide on the volume of whole blood, the scalingpoint, and the position of float; the focal point was to distinguish the information ofthe middle three layers in the irradiation of blue light source. Also the borders of themiddle three layers were determined by adopting mutual contrast of red picture andgreen picture, and the transformation of algorithm rules and routine blood parameterswere defined.
3) Research on the control module of sample in and out of capillaries, and theirdetection. The mechanical structure of sample in and out of capillaries has beendesigned with DSP as the CPU, which reduced electric circuits around. Experimentson the precision of sample in and out of capillaries and detection have been performed,indicating a good stability of the system.
4) A rapid blood test system on the basis of QBC technique was constructed bycombining the new test method with some techniques such as optical path and electriccircuit. This paper gave a detailed description of the three components of the systemas detection structure of optical path, control structure of sample in and out ofcapillaries and data processing algorithm.
5) A medical experiment has been designed and conducted to test the feasibilityand reliability of the system. In contrast with QBC Autoread plus and SysmexXT-1800i with HCT, HGB, GRAN, LM, WBC and PLT as the contrast parameters, asatisfactory result has been made in terms of their correlation coefficients, which were0.99,0.98,0.95,0.93,0.96,0.94and0.99,0.97,0.986,0.95,0.975,0.94respectively.
Result: A rapid blood cells detection method was proposed on the basis of dry-type chemical method and a rapid blood detection system based on QBC technique wasconstructed with DSP as the center of the system and by integrating with optical pathdetection techniques and other techniques including electric circuit control and dataprocessing. A set of model machine was made to perform on it experimental test of alot of samples, by which the feasibility and reliability of the system was verified. Onenational patents of invention has been applied for sample in-and-out of capillarysystem.
Conclusion: This system is a breakthrough of the traditional blood detectionconception and realization of blood detection on dry-type chemical method basis.Compared with wet-type detection method, this system is more flexible and free ofliquid reagent treatment. This project has accomplished the better functions of moreportable, vibration resistant and test results more rapid and accurate on fieldconditions. A timely and accurate test results will save much time in rescuing thepatients and a more compact construction and less operating cost will be moresuitable to be used in basic-level medical units. A rapid blood detection system basedon QBC technology has some advantages as below: firstly it has high performanceand is convenient to use; secondly, it can accomplish the quantitative measurement ofsuch nine parameters of blood cells as HCT, HGB, RBC, PLT, MCHC,GRAN,%GRAN, LM and%LM, and it can also perform such functions asself-caliberation, warning, output storage and printing. So it could find a widerapplication in rapid field blood detection and basic-level emergency call, and it is ofirreplaceable significance especially when it is used to perform routine hemogramdetection in war time.
Characteristics and innovations:
1) Centrifugation and detection are performed separately. Except for the fiveminutes spent on centrifugation, it takes60seconds to finish a detection of aspecimen, which greatly improves the speed of detection. Especially when there aremany specimens, taking20specimens for instance, it takes25minutes for this systemto finish their detection (including5minutes spent on centrifugation), whereas it takesQBC Autoread plus45minutes and QBC Star120minutes.
2) The detection precision has been improved through double light sourcesdetection, axial rotation of capillaries, and instant photography of high resolutionimage sensor of color linear array CCD.
3) Cold light source LED has low power consumption, wide range of working temperatures, good environmental suitability, and strong vibration resistance andanti-interference. Also it is long-lived and easy to control.
4) The adoption of DSP as the core controller can achieve the processing of allkinds of digital signals rapidly and at real time. In terms of software, the control oftwo motors has been scheduled in the form of tasks so as to increase the responsespeed of the system.
5) The module of detection has independent function and greatly simplifiedstructure, so it is small sized, light in weight and easy to carry around. In this sense, itis superior to similar products at home and abroad in that it has effectively broughtdown the cost and improved its integration.
方法：首先，在研究总结了前人的研究成果基础上提出血细胞分析检测方法：离心、检测相分离的基于血液棕黄色层定量离心分层技术（QBC）的方法。为离心机设计可快速拆卸的专用离心盘，用来配合干式血液检测毛细管的离心，而在更换离心盘后，该离心机可用于常规的临床检验实验室用途，这样可以做到一机多用。该算法对传统检测方法进行了改进：首先，采用彩色线阵CCD结合自行设计的成像透镜对待测毛细管实现一次成像，大大提高检测精度与速度；其次，在用蓝光激发荧光照射时，使毛细管沿其轴向转动10个位置检测取平均值，减小了由于离心过程中各个界面分层不均带来的影响；最后，通过医学实验对系统的可靠性和可行性进行了检验分析。
内容：本论文主要包括以下几个方面的研究工作：
（1）进行了光路及CCD图像采集模块的研究。在光路及CCD图像采集模块中，采用LED冷光源增强了环境适应性；应用彩色线阵CCD不需额外的滤光片，在信号处理电路中采用相关双采样法及视频信号处理专用芯片AD9826，简化了系统的结构；应用CPLD可编程逻辑器件实现CCD驱动的时序，简化程序，提高应用性。
（2）对图像采集后的信号处理算法进行了研究。该部分主要是确定各层分层的确切位置，在红光源照射下，确定全血体积、标定点及浮子位置；在蓝光源照射下，重点区分中间三层（白细胞和血小板层）的信息。中间三层的边界确定采用红色图和绿色图相互比照的方法。确定了算法规则及血常规参数的转化。
（3）对系统毛细管进排样、检测控制模块进行了研究。设计了毛细管进排样的机械结构，中央处理单元采用DSP数字信号处理器，减少外围电路，进行了进排样精度及检测精度的实验，系统稳定性良好。
（4）在研究的新型检测方法基础上，结合光路技术和电路技术等，构建了“基于定量离心分层技术的快速血液检测系统”。本文详细叙述了系统三个组成部分：光路检测结构、毛细管进排样结构及数据处理算法的实现。
（5）设计并开展了一项医学实验，对系统的可靠性和可行性进行检验。在与QBC Autoread plus和Sysmex XT-1800i两类仪器的对比中，以红细胞压积（HCT）、血红蛋白（HGB）、粒细胞（GRAN）、淋巴细胞及单核细胞（LM）、白细胞（WBC）和血小板（PLT）为对比参数，相关系数分别为0.99、0.98、0.95、0.93、0.96、0.94和0.99、0.97、0.986、0.95、0.975、0.94，达到了令人满意的效果。
结果：
提出了基于干式化学法的快速血细胞检测方法，并在以数字信号处理器（DSP）为核心控制的系统中，结合光路检测技术、电路控制技术和数据处理技术，构建了“基于定量离心分层技术的快速血液检测系统”检验平台；制作完成实验样机一套，通过对大量样本的实验测试，验证了系统的可靠性和可行性。针对基于干式化学法的血液检测系统的毛细管进排样系统，申请发明专利一项。
结论：
本系统完全突破了传统的血液检测概念，实现了基于干式化学法的检测，相比湿式检测具有灵活、不需要液体试剂处理过程。本课题实现了现场条件下仪器携行性、抗震性好、检测结果快速准确的功能。一方面及时准确的检测结果为伤病员的抢救赢得了时间；另一方面，系统结构紧凑、运营成本低，同样适合基层医疗单位使用。“基于定量离心分层技术的快速血液检测系统”性能优良、使用方便，运用该系统可实现血细胞HCT、HGB、RBC、PLT、MCHC、GRAN、%GRAN、LM和%LM9个参数的定量测量，并具有自校准和报警提示、输出存储与打印等功能。在现场血液快速检测和基层急诊中有较广泛的应用价值，特别是在战时状态进行常规血细胞检测具有不可替代的价值。
研究意义：
1）样本无需制备，可迅速展收，适合现场快速检验；
2）检测速度快，除去离心时间（5分钟），进样、检测时间≯60s，操作简便，无需复杂维护，携行性好；
3）结构紧凑，体积小，环境适应性强，使用综合成本低。
本研究特色及创新之处：
1）离心、检测分离，除去离心的5分钟，每个样本检测时间在60秒内，大大提高检测速度，尤其提高多样本的检测速度（以20样本为例，本系统与QBCAutoread plus和QBC star相比，三者检测时间分别是25分钟、45分钟和120分钟）；
2）双光源检测、毛细管轴向转动，高分辨率的彩色线阵CCD图像传感器一次成像，提高检测精度；
3）冷光源LED功耗低、控制简单、寿命长，工作温度范围广，环境适应性、抗震动和抗干扰能力强；
4）采用DSP作为核心控制器，实时、快速地实现各种数字信号的处理，软件方面把对两个电机的控制以任务的形式进行调度，提高了系统的反应速度；
5）检测模块功能独立，结构大大简化，体积小，重量轻，携行性好，有效降低成本，提高可集成性，优于国内外的同类产品。
Objective: The contents of various blood cells in the blood are important parametersto reflect the health of human body. The blood flows through all the tissues andorgans and any change of its contents will exercise great influence on the realizationof the functions of human bodies and tissues. Likewise, any pathological change inthe functions will also influence blood contents. So analysis of the blood cells inhuman bodies is of great importance. Currently, clinical detection equipments, whichare mainly based on wet-type chemical method, have some disadvantages in commonsuch as low speed of detection, big size, requiring large amount of liquid reagents togo with, restriction in field application, and so on. This paper aimed to research thekey technology and equipments of rapid blood detection based on QBC technique, adry-type chemical method, so as to achieve the quantitative detection of normal cellparameters and find its application in field occasion, emergency situation andbasic-level medical units.
Methods: Based on the research and summary of achievements previously made, anew method to detect blood cells was proposed as QBC technique which realized theseparation of centrifugation and detection on the basis of quantitative buffy coat. Acentrifugal disc, which could be disassembled very conveniently and quickly, wasdesigned specifically for the centrifuge, which could also be used to routine clinicallaboratory when the special centrifugal disc was replaced so as to realize more thanone purpose in a machine. The method has made some improvements on thetraditional test methods as follows: firstly, color linear array CCD, combined withself-designed imaging lens was adopted to fulfill instant photograph of theto-be-tested capillaries, which greatly improved the accuracy and speed of detection;secondly, the capillaries were designed to rotate along the axial direction when theywere irradiated with fluorescence excited by blue lights, then they were detected at10times to get their average value, as reduced the effect brought about by unevenness ofthe layering of interfaces in the course of centrifugation; finally the system was examined to prove its feasibility and reliability through medical experiments.
Contents: This paper is mainly composed of the researches in the following aspects:
1) Research on optical path and CCD image collection module. In the module ofoptical path and image collection, LED cold light source was employed to enhance itsenvironmental suitability; the configuration of the system was simplified by usingcolor linear array CCD to replace additional spectral filter, and by adopting coupledouble sample (CDS) method and AD9826, a special-purpose video-signal processingchip in the signal processing electric circuit; the application of CPLD, aprogrammable logic device, was the time sequence to realize the drive of CCD, whichcould simplify procedures and improve the applicability.
2) Research on the signal processing algorithm after the collection of images.This part was intended to determine the definite position of each layer, and in theirradiation of red light source, to decide on the volume of whole blood, the scalingpoint, and the position of float; the focal point was to distinguish the information ofthe middle three layers in the irradiation of blue light source. Also the borders of themiddle three layers were determined by adopting mutual contrast of red picture andgreen picture, and the transformation of algorithm rules and routine blood parameterswere defined.
3) Research on the control module of sample in and out of capillaries, and theirdetection. The mechanical structure of sample in and out of capillaries has beendesigned with DSP as the CPU, which reduced electric circuits around. Experimentson the precision of sample in and out of capillaries and detection have been performed,indicating a good stability of the system.
4) A rapid blood test system on the basis of QBC technique was constructed bycombining the new test method with some techniques such as optical path and electriccircuit. This paper gave a detailed description of the three components of the systemas detection structure of optical path, control structure of sample in and out ofcapillaries and data processing algorithm.
5) A medical experiment has been designed and conducted to test the feasibilityand reliability of the system. In contrast with QBC Autoread plus and SysmexXT-1800i with HCT, HGB, GRAN, LM, WBC and PLT as the contrast parameters, asatisfactory result has been made in terms of their correlation coefficients, which were0.99,0.98,0.95,0.93,0.96,0.94and0.99,0.97,0.986,0.95,0.975,0.94respectively.
Result: A rapid blood cells detection method was proposed on the basis of dry-type chemical method and a rapid blood detection system based on QBC technique wasconstructed with DSP as the center of the system and by integrating with optical pathdetection techniques and other techniques including electric circuit control and dataprocessing. A set of model machine was made to perform on it experimental test of alot of samples, by which the feasibility and reliability of the system was verified. Onenational patents of invention has been applied for sample in-and-out of capillarysystem.
Conclusion: This system is a breakthrough of the traditional blood detectionconception and realization of blood detection on dry-type chemical method basis.Compared with wet-type detection method, this system is more flexible and free ofliquid reagent treatment. This project has accomplished the better functions of moreportable, vibration resistant and test results more rapid and accurate on fieldconditions. A timely and accurate test results will save much time in rescuing thepatients and a more compact construction and less operating cost will be moresuitable to be used in basic-level medical units. A rapid blood detection system basedon QBC technology has some advantages as below: firstly it has high performanceand is convenient to use; secondly, it can accomplish the quantitative measurement ofsuch nine parameters of blood cells as HCT, HGB, RBC, PLT, MCHC,GRAN,%GRAN, LM and%LM, and it can also perform such functions asself-caliberation, warning, output storage and printing. So it could find a widerapplication in rapid field blood detection and basic-level emergency call, and it is ofirreplaceable significance especially when it is used to perform routine hemogramdetection in war time.
Characteristics and innovations:
1) Centrifugation and detection are performed separately. Except for the fiveminutes spent on centrifugation, it takes60seconds to finish a detection of aspecimen, which greatly improves the speed of detection. Especially when there aremany specimens, taking20specimens for instance, it takes25minutes for this systemto finish their detection (including5minutes spent on centrifugation), whereas it takesQBC Autoread plus45minutes and QBC Star120minutes.
2) The detection precision has been improved through double light sourcesdetection, axial rotation of capillaries, and instant photography of high resolutionimage sensor of color linear array CCD.
3) Cold light source LED has low power consumption, wide range of working temperatures, good environmental suitability, and strong vibration resistance andanti-interference. Also it is long-lived and easy to control.
4) The adoption of DSP as the core controller can achieve the processing of allkinds of digital signals rapidly and at real time. In terms of software, the control oftwo motors has been scheduled in the form of tasks so as to increase the responsespeed of the system.
5) The module of detection has independent function and greatly simplifiedstructure, so it is small sized, light in weight and easy to carry around. In this sense, itis superior to similar products at home and abroad in that it has effectively broughtdown the cost and improved its integration.
引文
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