Electrochemiluminescence method of detecting an analyte in a liquid sample and analysis system

Abstract

An electrochemiluminescence method of detecting an analyte in a liquid sample and a corresponding analysis system. An analyte in a liquid sample is detected by first providing a receptacle containing a fluid comprising protein coated magnetic microparticles to a stirring unit. Stirring of the fluid is necessary since the density of the microparticles is usually higher than the density of the buffer fluid. Thus the microparticles tend to deposit on the bottom of the receptacle leading to an aggregation of the microparticles because of weak interactions. To obtain representative measurements a homogeneous distribution of the microparticles in the buffer fluid is necessary to ensure a constant concentration of microparticles for each analysis cycle. It is further necessary to provide disaggregation of the microparticles, which is also realized by stirring the fluid. Stirring is conducted with a rotational frequency that is adapted to the amount of fluid to be stirred.

What is claimed is: 
     
         1 . An electrochemiluminescence method of detecting an analyte in a liquid sample using a series of analysis cycles, the method comprising, for each analysis cycle in the series of analysis cycles:
 providing a receptacle containing a fluid comprising protein coated magnetic microparticles to a stirring unit,   acquiring a signal being indicative of an amount of the fluid contained in the receptacle,   determining a rotational frequency for the stirring unit dependent on the amount of fluid in the receptacle, the rotational frequency being proportional to the amount of fluid,   stirring the fluid for a predefined period of time by applying the previously determined rotational frequency, the predefined period of time being held constant for each stirring process, wherein the stirring is performed for homogenization and disaggregation of the microparticles,   extracting a portion of the fluid comprising the protein coated magnetic microparticles from the receptacle, thereby reducing the amount of the fluid contained in the receptacle,   mixing a portion of the liquid sample with the portion of the fluid comprising the protein coated magnetic microparticles and with a marker,   incubating the mixture comprising the analyte, the protein coated magnetic microparticles, and the marker in an incubator,   transporting a portion of the mixture from the incubator to a measurement cell,   applying a magnetic field to the measurement cell for magnetic adhesion of the protein coated magnetic microparticles to a working electrode of the measurement cell,   applying an excitation energy for causing luminescence,   measuring of the luminescence for acquisition of a measurement signal, and   generating an output signal being indicative of the presence of the analyte in the liquid sample using the measurement signal   
       wherein the rotational frequency of the stirring unit is maximally high but below a frequency that causes foaming. 
     
     
         2 . The method of  claim 1 , wherein extracting a portion of fluid from the receptacle employs a pipetting probe, and wherein acquiring a signal being indicative of the amount of fluid contained in the receptacle is conducted using the pipetting probe. 
     
     
         3 . The method of  claim 1 , wherein acquiring a signal being indicative of the amount of fluid contained in the receptacle employs a capacitive method. 
     
     
         4 . The method of  claim 1 , wherein determining the rotational frequency for the stirring unit comprises:
 providing a first signal being indicative of the amount of fluid contained in the receptacle to a data processing unit,   in response to providing the first signal to the data processing unit receiving a second signal being indicative of the rotational frequency appropriate for the amount of fluid from the data processing unit.   
     
     
         5 . The method of  claim 4 , wherein the data processing unit comprises a table, the table comprising information indicative of which rotational frequency to use for a defined amount of fluid, wherein the data processing unit in response to receiving the first signal looks up the table for the rotational frequency appropriate for the amount of fluid the first signal is indicative of and provides a second signal being indicative of the rotational frequency appropriate for the amount of fluid. 
     
     
         6 . The method of  claim 5 , wherein the data comprised in the table is determined empirically. 
     
     
         7 . The method of  claim 1 , wherein stirring is only conducted prior to extracting fluid from the receptacle. 
     
     
         8 . The method of  claim 1 , wherein the excitation energy is applied by applying electrical energy, radiation energy and/or chemical energy. 
     
     
         9 . The method of  claim 1 , the marker being capable of effecting electrochemiluminescence, wherein an electrochemically active substance contributes to an electrochemiluminescence reaction with the marker resulting in the luminescence. 
     
     
         10 . The method of  claim 8 , further comprising:
 carrying out a reaction sequence comprising at least one specific biochemical binding reaction to form a complex as a result of the presence of the analyte in the liquid sample, the complex comprising the marker and said complex further being bound to a magnetic microparticle,   carrying out a detection cycle in a measuring cell having a working electrode for determination of the presence of said analyte, said detection cycle comprising a capturing step during which the complex is contacted with a working electrode in such a manner that said microparticle is attracted by the magnetic field of a magnetic component positioned on the side of the working electrode facing away from the sample, thereby being deposited on the surface of said working electrode facing the sample, and applying a potential to the working electrode to trigger the electrochemiluminescence reaction of the marker with the electrochemically active substance causing the luminescence of the marker to thereby determine the presence of the analyte in the liquid sample.   
     
     
         11 . The method of  claim 1 , wherein the luminescence is measured using an optical sensor, and wherein the measurement signal is acquired by sampling an output signal of the optical sensor. 
     
     
         12 . An electrochemiluminescence analysis system for detecting an analyte in a liquid sample using a series of analysis cycles, the system comprising:
 a stirring unit for stirring a fluid comprising protein coated magnetic microparticles provided in a receptacle,   a measuring unit being operable to generate a signal indicative of an amount of fluid in the receptacle,   an extraction component for extracting a portion of the fluid comprising the protein coated magnetic microparticles from the receptacle,   an incubator for receiving a liquid comprising the analyte, the portion of magnetic microparticles and a marker for marking the analyte, the marker being capable of effecting luminescence upon application of excitation energy,   a trigger component for applying the excitation energy for causing the luminescence,   an acquisition component for measuring the luminescence the acquisition component being operable to provide a measurement signal,   a data processing unit being adapted to:
 determine a rotational frequency for the stirring unit using the signal indicative of the amount of fluid in the receptacle, the rotational frequency being proportional to the amount of fluid, 
 control the stirring unit to stir the fluid for a predefined period of time by applying the previously determined rotational frequency, the predefined period of time being held constant for each stirring process, wherein the stirring is performed for homogenization and disaggregation of the microparticles, wherein the rotational frequency of the stirring unit is maximally high but below a frequency that causes foaming, 
 generate an output signal being indicative of the presence of the analyte in the liquid sample using the measurement signal. 
   
     
     
         13 . The analysis system of  claim 12  further comprising a database, the database comprising a table comprising information indicative of which rotational frequency to use for a defined amount of fluid, wherein the data processing unit is further operable to access the database and look up table for the rotational frequency appropriate for the amount of fluid the first signal is indicative of. 
     
     
         14 . The method of  claim 2 , wherein acquiring a signal being indicative of the amount of fluid contained in the receptacle employs a capacitive method. 
     
     
         15 . The method of  claim 2 , wherein determining the rotational frequency for the stirring unit comprises:
 providing a first signal being indicative of the amount of fluid contained in the receptacle to a data processing unit,   in response to providing the first signal to the data processing unit receiving a second signal being indicative of the rotational frequency appropriate for the amount of fluid from the data processing unit.   
     
     
         16 . The method of  claim 2 , wherein stirring is only conducted prior to extracting fluid from the receptacle. 
     
     
         17 . The method of  claim 2 , wherein the excitation energy is applied by applying electrical energy, radiation energy and/or chemical energy. 
     
     
         18 . The method of  claim 2 , the marker being capable of effecting electrochemiluminescence, wherein an electrochemically active substance contributes to an electrochemiluminescence reaction with the marker resulting in the luminescence. 
     
     
         19 . The method of  claim 2 , wherein the luminescence is measured using an optical sensor, and wherein the measurement signal is acquired by sampling an output signal of the optical sensor.