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WO1992021973A1 - Procede et dispositif de detection pour determiner des marqueurs de l'infarctus du myocarde - Google Patents

Procede et dispositif de detection pour determiner des marqueurs de l'infarctus du myocarde Download PDF

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WO1992021973A1
WO1992021973A1 PCT/SE1992/000386 SE9200386W WO9221973A1 WO 1992021973 A1 WO1992021973 A1 WO 1992021973A1 SE 9200386 W SE9200386 W SE 9200386W WO 9221973 A1 WO9221973 A1 WO 9221973A1
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Prior art keywords
analytes
ligand
analyte
sample
sensor
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PCT/SE1992/000386
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English (en)
Inventor
Lennart Pedersen
Ralph STÅLBERG
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Biacore AB
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Pharmacia Biosensor AB
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Priority to AU19756/92A priority Critical patent/AU656202B2/en
Priority to JP4511118A priority patent/JPH06508204A/ja
Publication of WO1992021973A1 publication Critical patent/WO1992021973A1/fr
Priority to PCT/SE1993/000488 priority patent/WO1993025910A1/fr
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

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    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/17—Systems in which incident light is modified in accordance with the properties of the material investigated
    • G01N21/55—Specular reflectivity
    • G01N21/552—Attenuated total reflection
    • G01N21/553—Attenuated total reflection and using surface plasmons
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/543—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals
    • G01N33/54366—Apparatus specially adapted for solid-phase testing
    • G01N33/54373—Apparatus specially adapted for solid-phase testing involving physiochemical end-point determination, e.g. wave-guides, FETS, gratings
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/573—Immunoassay; Biospecific binding assay; Materials therefor for enzymes or isoenzymes
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
    • G01N33/6893—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids related to diseases not provided for elsewhere
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2800/00—Detection or diagnosis of diseases
    • G01N2800/32—Cardiovascular disorders
    • G01N2800/324—Coronary artery diseases, e.g. angina pectoris, myocardial infarction

Definitions

  • This invention relates to a method of simultaneously assessing from blood samples the variation with time of at least two different and independent markers indicative of myocardial damage to provide for early and reliable diagnosis of myocardial infarction as well as to improve the monitoring of the treatment and recovery after a myocardial event.
  • the invention also relates to a sensor unit to be used in the method.
  • Acute myocardial infarction is a major cause of deaths in the developed countries, the medical term describing the development of ischemia and necrosis of a portion of the myocardium.
  • the ischemia is caused by occlusion of the artery by a thrombus preventing normal blood flow to the affected area of the myocardium and eventually resulting in tissue necrosis.
  • the current clinical diagnosis of AMI is based on chest pain (sudden onset of pain which sustains for more than 15 minutes) , electrocardiogram (ECG) and levels and kinetics of heart muscle specific enzymes and proteins.
  • the treatment with thrombolytics has severe side effects such as stroke and haemorrhagic bleedings.
  • tissue plasminogen activator (tPA) and streptokinase are hit with these side effects.
  • Accurate diagnosis is thus fundamental in these cases.
  • the great economic importance of such early identification of the patients not requiring the AMI treatment is readily appreciated, considering the substantial cost reductions obtained when such patients after a much shorter time than before may be transferred to an ordinary medical ward or in the most favourable cases can be sent home.
  • myosin light chain Another protein known to be released from cardiac muscle following myocardial infarction and forming the basis of a test is myosin light chain.
  • Each one of the mentioned marker proteins exhibits specific serum concentration changes corresponding to the kinetics and release of the respective protein in an acute infarction.
  • WO91/01498 discloses the simultaneous or sequential testing for creatine kinase and myosin light chain for the early detection of a myocardial infarction. While the combination of the two assays is said to provide a very reliable diagnostic test for the early phase of a heart attack, it also permits differentiation between a myocardial infarction and other ischemic events causing cardiac pain, such as angina pectoris. The latter feature is due to the fact that myosin light chain is released not only in case of an acute myocardial infarction, but also for other cardiac injuries, whereas creatine kinase is substantially only released in the instance of an acute myocardial infarction.
  • One object of the present invention is therefore to provide an assay method by which it is possible to reliably diagnose AMI in such a short time that a treatment with thrombolytics will be beneficial.
  • Another object of the invention is to provide an assay method for the diagnosis of AMI or monitoring of treatment with thrombolytics that may be performed at or close to the patient's bedside.
  • Still another object of the invention is to provide an assay method for monitoring cardiac enzymes to detect the possible initiation of an AMI during thorax surgery, and which may be performed in the operating room.
  • a further object of the invention is to provide a sensor means for use in the assay methods mentioned above.
  • the above mentioned objects are achieved with the method and sensor means of the present invention.
  • a basic concept of the invention resides in determining with short intervals the variation with time of at least two, preferably at least three different markers, i.e. enzymes or proteins, of myocardial infarction in real-time measurements in a flow cell system, which measurements are based upon interactions of the analytes with ligands bound to a sensor surface, the interactions causing a detectable change of the physico-chemical characteristics of the surface.
  • CK-MM A and CK-MM B are not different analytes in the context of the present invention.
  • a method of AMI marker determination or monitoring comprising the steps of (i) simultaneously determining from a first blood, serum or plasma sample from a patient at least two, preferably at least three different analytes indicative of myocardial infarction by contacting in a flow cell or cells the sample with one or more sensor surface areas each supporting a different ligand or mixture of different ligands capable of specifically binding to a respective analyte, and, optionally after additionally binding an analyte specific reagent or reagent complex to the bound analytes, detecting any binding " interaction of each analyte with its ligand as a consequential change of the physico- chemical characteristics of the sensor surface;
  • step (iii) repeating step (i) for at least a second blood, serum or plasma sample taken from the patient at a determined time interval from said first sample;
  • reagent complex as used herein is meant that the reagent is bound to one ore more other species. Such a reagent complex may be added as an assembly or be formed successively after the reagent has bound to the analyte as will be further elucidated below.
  • a sensor means comprising, immobilized to one or more sensing areas thereof, either individually or in combination, at least two, preferably at least three different ligands, each ligand being capable of specifically binding to a respective analyte indicative of myocardial infarction, said sensor means being adapted for the detection of any analyte-ligand interaction as a consequential change of the physico-chemical characteristics of the sensing surface, and said ligand supporting surface areas being regeneratable after the coupling of analytes thereto.
  • the different ligands may thus be either individually immobilized to respective sensing areas, or co-immobilized to a single sensing area. Combinations of these embodiments are, of course, also possible, i.e. that there are two or more sensing areas, each area supporting co-immobilized ligands.
  • the sensor surface is a surface capable of exhibiting surface plasmon resonance (SPR) and the detection of the cardiac analytes or markers is carried out by surface plasmon resonance spectrometry.
  • SPR surface plasmon resonance
  • Fig. 1 is a schematic illustration of a per se known SPR-based flow cell measurement system
  • Fig. 2 is an exploded sectional partial view of a flow cell unit useful for the purposes of the present invention
  • Fig. 3 is an SPR-sensor diagram showing the co- immobilization of a monoclonal antibody specific for CK-MB and a monoclonal antibody specific for myoglobin to a sensing surface;
  • Fig. 4 is a corresponding diagram as in Fig. 3 showing the analysis of a sample containing elevated levels of CK- MB and myoglobin using the sensing surface with immobilized CK-MB and myoglobin monoclonals in Fig. 3;
  • Fig. 5 is a corresponding diagram as in Fig. 4 showing the analysis of a sample not containing CK-MB and only a normal level of myoglobin.
  • one basic feature of the invention resides in the simultaneous measurement of several, viz. at least two, preferably, however, at least three, or even e.g. four or five, different myocardial infarction markers or analytes. While the diagnostic sensitivity and specificity of such markers will vary, the appropriate selection of analytes and the determination thereof simultaneously from blood samples taken at short intervals will provide for an analytical picture or pattern which will be very useful for diagnosing or excluding AMI. The relationship between the various analytes during a certain time interval will also give an indication of how long an infarction condition has been going on, for example based upon the rise of the level of one analyte and the decline of another. Likewise, the areas under the respective analyte graphs will indicate the extent of the damage to the cardiac muscle tissue.
  • Another basic feature of the invention lies in the already mentioned repeated analyses at, preferably, regular short intervals permitting the variation of time of the cardiac markers to be determined. Such information will considerably improve the possibilities of making a reliable AMI diagnosis or exclusion.
  • An apparatus system enabling the desired type of analytical procedures to be performed should (i) rely on a measuring principle based upon the interaction of the analytes with ligands immobilized to a sensor surface and detecting complex formation as a consequential change of the physico-chemical properties of the sensor surface; (ii) have a flow cell system permitting the desired simultaneous detection of several analytes; and (iii) have a sensor surface or surfaces which can be regenerated in situ in the sense that bound analytes may be removed from the respective ligands to permit consecutive analyses on one and the same sensor surface(s) after a regenerating step. All the requirements (i) to (iii) may be met by analytical technology which is known per se in the art.
  • further complexing of the bound analytes may be accomplished by reacting the ligand bound analytes with a secondary reagent specific to .the analyte, i.e. a sandwich assay, as is per se known in the art.
  • a secondary reagent specific to .the analyte, i.e. a sandwich assay, as is per se known in the art.
  • Such bound secondary reagent may then, if desired, be further complexed by reacting it with a tertiary reagent, etc. to still more increase the surface change to be detected.
  • optically dense species such as a particulate label, for example, glass, latex, colloidal silver or gold, metal oxide or ferritin; see e.g. EP-A-276 142.
  • the measuring principle as defined above will enable real-time measurements to be conducted and thereby permit the desired performance of consecutive analyses of blood samples taken with short intervals.
  • a measuring principle, and the sensor surface associated therewith is also readily adaptable to flow cell systems.
  • internal reflectance methods particularly evanescent wave spectroscopy (EWS) , such as attenuated internal reflection (ATR) spectroscopy, total internal reflectance fluorescence (TIRF) spectroscopy, and surface plasmon resonance (SPR) spectroscopy, or wave guide spectroscopy. All these methods are based upon the examination of an optical property of a solution bordering a surface where total internal reflection has occurred.
  • EWS evanescent wave spectroscopy
  • ATR attenuated internal reflection
  • TIRF total internal reflectance fluorescence
  • SPR surface plasmon resonance
  • sensors that may be used are those based upon photoacoustic piezoelectric, surface acoustic wave (SAW) , or electrochemical measurements, etc. Particularly suitable for the purposes of the invention is SPR spectroscopy as will be described in more detail below.
  • SAW surface acoustic wave
  • a flow cell system is essential for providing a rapid and easy to handle system apt to automation.
  • flow cell (which is to be interpreted in a broad sense) is meant that the sample and other analytical fluids will flow past the sensing surface or surfaces at a constant rate.
  • Such a system may, as mentioned above, in one embodiment thereof comprise a plurality of sensing surface areas, i.e. one for each cardiac analyte as well as a control area, and optionally one or more areas for other purposes as will be described below.
  • the flow may be either in parallel or in series with respect to the different sensing surface areas.
  • a parallel flow cell system may comprise several separate flow cells arranged in parallel, whereas in a series system several separate flow cells may be connected in series.
  • a parallel or serial flow system may also be formed by several defined sensing surface areas contained within a single flow cell.
  • a flow cell system comprising several flow cells
  • the ligands for all the cardiac analytes may, as already mentioned above, be co-immobilized in a single surface area.
  • the different immobilized capturing molecules are immobilized to different detection areas as described above.
  • the independence of area size for the. surface concentration detecting device will, however, in accordance with this novel and inventive concept, allow reduction in the number of detection areas by co- immobilization of two or more different capturing molecules onto the same area, the specificity then being revealed by sequential injections of second reagents (such as antibodies) specific for each analyte bound by the respective capturing molecules.
  • second reagents such as antibodies
  • Sensor surfaces which may be regenerated as defined above a substantial number of times, e.g. 50 to 100 times, are known per se in the art and are, for example, described in our published PCT-application WO 90/05305 (the full disclosure of which is incorporated by reference herein) .
  • These sensor surfaces comprise a substrate coated with a metal film to which has been attached a layer of an organic polymer or a hydrogel forming a so-called basal surface which may contain functional groups for selectively binding the desired ligands.
  • W0 90/05303 the full disclosure of which is incorporated by reference herein
  • Such a surface is easily regeneratable in situ in a flow cell.
  • the bound analytes can be removed from the respective ligands to prepare the sensing surface(s) for a subsequent assay of another sample. This is made possible by the binding of the analyte to the ligand being broken by the regenerating fluid whereas the binding of the ligand to the sensor surface is not.
  • myocardial infarction analytes or markers from which the test panel may be selected are myoglobin, creatine kinase (CK) (also known as creatine phosphokinase) and its isoenzymes and isomers thereof, especially CK-MB, tropomyosin, lactate dehydrogenase (LD) , troponin C, T and I, aspartate aminotransferase, and myosin, especially the light chain thereof.
  • CK creatine kinase
  • LD lactate dehydrogenase
  • troponin C troponin C
  • T and I aspartate aminotransferase
  • myosin especially the light chain thereof.
  • CK-MB CK-MB
  • myoglobin myoglobin
  • troponin I CK-MB
  • none of the above mentioned markers is absolutely specific for acute cardiac muscle damage.
  • both CK-MB and myoglobin may also be present in skeletal muscle damages.
  • One example of such a control analyte is carbonic anhydrase III (CA III) which has been shown to be present in substantial amounts in skeletal muscle, in minor amounts in i.a. smooth muscle cells, but not at all in the myocardium.
  • CA III can therefore be used to distinguish whether increased concentrations of e.g. myoglobin originate from cardiac or skeletal muscle (see e.g. Vaananen H.K. et al. , Clin. Chem. 36/4, 635-638 (1990) .
  • the ligands for the specifically binding cardiac analytes are usually antibodies, particularly monoclonal antibodies.
  • antibodies as used herein is also to be understood active antibody fragments, antibodies and fragments thereof produced by genetic engineering, etc.
  • the functionalizing of the basal sensor surface with the ligands is simplified if the ligands are chimeric molecules, i.e. comprise a common part for binding to the basal surface and a variable part for binding to the different cardiac analytes.
  • the sensing surface comprises a basal dextran layer
  • the chimeric molecule may consist of an antibody to dextran which is conjugated to a monoclonal directed against the desired cardiac analyte.
  • Antibodies that may be used as ligands are described in the prior art, and may also be produced by methods known per se, e.g. by hybridoma or recombinant DNA technology.
  • antibodies against creatine kinase and creatine kinase MB are, for example, described in EP-A-288 179, EP- A-339 814, EP-A-261 781, and US-A-4,912,033.
  • Antibodies against myosin light chain are, for instance, disclosed by WO 91/01498, WO 90/15329, and US-A- 4,879,216.
  • Antibodies directed against troponin T are described in, for example, EP-A-394,819, and antibodies against troponin I are described by Cummins B. et al., Bioehem. Soc. Trans. 15 (1987) 1060-61.
  • Myoglobin antibodies are, for instance, disclosed in JP-A-54011231.
  • Antibodies directed against lactate dehydrogenase are described in, for example, DE-A-2 350 711.
  • the blood samples taken from the patient may be analyzed directly as whole blood, but it may be preferable to use plasma or serum prepared from the blood sample, e.g. as obtained by an initial centrifugation or filtration step.
  • a further beneficial feature of the method of the present invention is that it may also provide information about the presence in the blood- samples of antibodies against streptokinase.
  • streptokinase is about tenfold cheaper than tissue plasminogen activator (tPA)
  • tPA tissue plasminogen activator
  • tPA is necessary if the patient has a high level of antibodies against streptokinase or is allergic thereto.
  • About 90% of the patients experiencing a reinfarcation within a year have neutralizing antibodies. Neutralizing antibodies can also occur after a flu, if the infection is caused by streptococcal bacteria.
  • the method of the invention preferably also comprises testing for such neutralizing streptokinase antibodies by including a streptokinase ligand (or an antibody against the anti- streptokinase antibody) in the or one of the sensing areas.
  • a streptokinase ligand or an antibody against the anti- streptokinase antibody
  • the present invention thus comprises determining (i) at least two, and preferably at least three, different infarction analytes; (II) one or optionally more control analytes; and (iii) streptokinase antibodies.
  • the angle of incidence of light directed towards an interface between two transparent media of different refractive indices exceeds a critical angle, the light is reflected back into the medium having the higher refractive index, so-called total internal reflection.
  • an electromagnetic field component of the light called the "evanescent wave” penetrates a short distance (of the order of a wave length) into the medium of lower refractive index.
  • the interface between the media is coated with a thin metal film, such as silver or gold, and the light is plane-polarized and monochromatic, the evanescent wave will at a certain angle of incidence interact with collective electron oscillations, called plasmons, in the metal.
  • SPR surface plasmon resonance
  • FIG. 1 A schematic illustration of an SPR based biosensor system of the flow cell type which is known in the art and may be used for the purposes of the invention is shown in Fig. 1.
  • a flow channel 1 has an open top portion covered by a sensor plate or chip 2 of glass coated with a metal film 3, more specifically of gold, to define a flow cell.
  • a prism 4 contacts the other side of the glass plate 2 to couple a wedge-shaped beam 5 of p-polarized light from a monochromatic light source 6 thereto.
  • the reflected light is directed against a detection unit 7 comprising a matrix (i.e. rows and columns) of photodetectors.
  • the gold film surface exposed to the fluid passing through flow cell 1 has ligands 8 immobilized thereto as will be further described below. Since the beam of light 5 reflected at the glass-metal interface represents a continuous range of incident angles, a shift in the resonance angle caused by a change in the concentration of biomolecules at the metal surface may be detected by the detector unit 7.
  • FIG. 2 schematically illustrates a part of a liquid handling block unit 9 comprising four flow cells 10, corresponding to flow cell 1 in Fig. 1.
  • the flow cells 10 are, as in Fig. 1, defined by upwardly open channel parts
  • sensor plate 12 covered by a sensor plate 12, corresponding to sensor plate 2 in Fig. 1.
  • optointerface 13 for effecting optical contact between the sensor plate 12 and a prism corresponding to prism 4 in Fig. 1.
  • the optointerface consists of a thin glass plate having elastic material pieces 14 on both sides with a refractive index matching that of the sensor plate 12.
  • the wedge-shaped beam 5 will extend transversely across the flow cells, each flow cell corresponding to, e.g., one column of photodetectors in the detector matrix 7.
  • the gold-coated glass plate 12 has, as a specific example known per se in the art, a hydrophilic matrix of non-crosslinked carboxymethylated dextran covalently bonded to the gold film through an optically and biologically inert linker layer of long chain hydrocarbon.
  • Ligands can be covalently immobilized to the dextran layer after activation, e.g. by derivatization with N-hydroxy- succinimide (NHS) , mediated by N-ethyl-N'-(dimethy1- aminopropyl)carbodiimide (EDC) .
  • NHS-ester formed readily reacts with uncharged primary amino groups of the ligands to be coupled thereto.
  • each flow cell may, for the purposes of the invention, in three of the flow cells 10 support ligands reactive with a respective cardiac analyte to be determined whereas the fourth flow cell preferably is used as a control or reference.
  • one flow cell may support a monoclonal antibody against creatine kinase MB (CK-MB) , a second flow cell a monoclonal antibody against myoglobin, a third flow cell a monoclonal antibody against troponin I, and a fourth flow cell a monoclonal antibody against carbonic anhydrase III (CA III) as a control.
  • CK-MB creatine kinase MB
  • CA III carbonic anhydrase III
  • more than four flow cells may be used if desired.
  • a fifth flow cell may support streptokinase ligands for determining streptokinase antibodies in a sample.
  • a sixth flow cell may support a ligand for an additional cardiac infarction analyte.
  • a test of a blood plasma sample for the cardiac enzymes creatine kinase MB, myoglobin, and troponin I using the analytical system illustrated in Figs. 1 and 2 may be performed as follows. After removal of the blood cells from a blood sample, a defined volume of the plasma is introduced into the liquid handling unit 9 and evenly distributed among the four flow cells 10. When flowing through the flow cells the mentioned cardiac enzymes, if present, will bind to the respective sensing surface supporting the proper ligand. This will cause a shift of the resonance angle, which shift is proportional to the amount of cardiac enzyme bound to the surface, as described above. Thereby the level of each analyte in the sample may be determined.
  • the ligand bound analytes are preferably reacted with a secondary reagent in sandwich assay fashion.
  • a secondary reagent may optionally be labelled with an optically dense species to still more increase the shift.
  • a tertiary reagent may be used.
  • the sensor surface is regenerated by passing a regenerating agent, e.g. glycine buffer, or phosphoric, formic or hydrochloric acid (10-100 mM) , through the cell. A whole such analytical procedure will take about 15 minutes.
  • a single flow cell may be used which has ligands for all the analytes of interest co-immobilized on the same sensing surface area. This embodiment will also be illustrated in the Example below.
  • the test panel of the sensor unit also contains a sensing surface area for testing for streptokinase antibodies, and information about the proper thrombolytic and dosage thereof to select may thereby be obtained at the same time, i.e. resulting in that the dose of streptokinase is adjusted or that tpA is selected rather than streptokinase.
  • the same bedside apparatus and test panel may then be used for monitoring the thrombolytic treatment to observe a reperfusion as soon as possible after it has taken place, so that the per se risky treatment may be stopped when no longer necessary or an alternative treatment may be introduced if reperfusion is not obtained in a reasonable time.
  • an analyte of interest e.g. CK-MB
  • relatively high levels of an analyte of interest e.g. CK-MB
  • the dynamic range of the secondary response may not be sufficient for permitting the development of the peak value of a specific analyte to be monitored.
  • the occurrence and exact monitoring of such a peak indicative of reperfusion may then be performed by instead measuring the primary response, i.e. the complex formation between the analyte and the immobilized ligand.
  • the above described procedure and method may also be used for monitoring the myocardium state during thorax surgery such that an AMI initiated during the surgery may be detected and treated before the thorax is closed.
  • EXAMPLE A Co-immobilization of monoclonal antibodies on sensing surface Immobilization of a monoclonal antibody specific for CK-MB and a monoclonal antibody specific for myoglobin was performed in the biosensor instrument in the following manner: A continuous flow of HBS (10 mM Hepes buffer, 0.15 M NaCl, 3.4 mM EDTA, 0.05 % Tween) , pH 7.4, over the sensing surface was maintained at 5 ⁇ l/min.
  • HBS 10 mM Hepes buffer, 0.15 M NaCl, 3.4 mM EDTA, 0.05 % Tween
  • Remaining reactive ester groups were deactivated by injection of 35 ⁇ l of 1 M ethanolamine hydrochloride, pH 8.5.
  • the sensorgram obtained is shown in Fig. 3 (response in resonance units, RU, plotted versus time in seconds) .
  • the response signal was evaluated at two levels: 20 seconds before the injection of EDC/NHS (A) and 9 minutes after the injection of ethanolamine (B) .
  • B minus A thus defines the immobilized amount of the two antibodies.
  • HBS 10 mM Hepes buffer, 0.15 M NaCl, 3.4 mM EDTA, 0.05 % Tween
  • 35 ⁇ l of a plasma sample containing CK-MB and myoglobin were injected into the instrument.
  • 4 ⁇ l each of second antibodies specific for CK-MB and myoglobin, respectively, at a concentration of 100 ⁇ g/ml were then injected in sequence followed by 4 ⁇ l of 10 mM glycine-HCl, pH 2.5.
  • the sensorgram obtained is shown in Fig. 4 (response in resonance units, RU, plotted versus time in seconds) .
  • the response signal was evaluated at four levels: 20 seconds before the injection of the sample (A) , 20 seconds before the injection of the second antibody specific for CK-MB (B) ,. 20 seconds before the injection of the second antibody specific for myoglobin (C) , and 20 seconds before the injection of glycine-HCl (D) .
  • A defines the baseline
  • B minus A defines the plasma response
  • C defines the specific response for CK-MB
  • D minus C defines the specific response for myoglobin.
  • the analysis time was 18 minutes.

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Abstract

Procédé de détermination des analytes résultant de l'infarctus du myocarde, comprenant les étapes suivantes: (i) détermination simultanée à partir d'un échantillon de sang, de sérum ou de plasma prélevé sur un patient d'au moins 2 différents analytes résultant de l'infarctus du myocarde, par mise en contact de l'échantillon avec la surface ou les surfaces (3) d'un détecteur cellulaire à passage de flux supportant des ligands (8) spécifiques de fixation des analytes, et détectant toute interaction de fixation sur la surface du détecteur (3); (ii) élimination de la surface du détecteur des analytes fixés; (iii) répétition des étapes (i)-(ii) pour au moins un second échantillon prélevé après un temps déterminé par rapport au premier échantillon; et (iv) détermination à partir des résultats obtenus de la variation dans le temps des analytes résultant d'une affection cardiaque. Un dispositif de détection (2, 3) destiné à être utilisé dans le procédé comprend, couplés de manière amovible à sa ou ses surfaces de détection (3), au moins deux différents ligands (8) spécifiques des analytes résultant de l'infarctus du myocarde.
PCT/SE1992/000386 1991-06-07 1992-06-05 Procede et dispositif de detection pour determiner des marqueurs de l'infarctus du myocarde Ceased WO1992021973A1 (fr)

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AU19756/92A AU656202B2 (en) 1991-06-07 1992-06-05 Method and sensor means for determining myocardial infarction markers
JP4511118A JPH06508204A (ja) 1991-06-07 1992-06-05 心筋梗塞マーカーの測定方法及びセンサー装置
PCT/SE1993/000488 WO1993025910A1 (fr) 1992-06-05 1993-06-02 Dosage de plusieurs analytes a l'aide de ligands co-immobilises

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SE9101735A SE9101735D0 (sv) 1991-06-07 1991-06-07 Method and sensor means for determining myocardial infarction markers
SE9101735-0 1991-07-07

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EP (1) EP0588891A1 (fr)
JP (1) JPH06508204A (fr)
AU (1) AU656202B2 (fr)
CA (1) CA2110705A1 (fr)
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WO (1) WO1992021973A1 (fr)

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WO2002023191A1 (fr) * 2000-09-12 2002-03-21 University Of Sydney Dosage diagnostique
WO2002057776A3 (fr) * 2001-01-19 2003-05-08 Quantech Ltd Systeme et procede pour reduire l'interference des particules lors de la detection d'analytes fluides
WO2003042669A1 (fr) * 2001-11-13 2003-05-22 Battelle Memorial Institute Spectroscopie photo-acoustique effectuee avec un reseau
US6673562B2 (en) 2000-08-24 2004-01-06 Spectral Diagnostics, Inc. Differential immunoassay
WO2004038415A1 (fr) * 2002-10-24 2004-05-06 Biacore Ab Test avec des ligands co-immobilises
EP1019717A4 (fr) * 1997-09-18 2004-09-01 Univ Utah Res Found Dispositif et procede de diagnostic
US6873415B2 (en) 2001-11-13 2005-03-29 Battelle Memorial Institute Photoacoustic spectroscopy sample array vessel and photoacoustic spectroscopy method for using the same
US6999174B2 (en) 2001-11-13 2006-02-14 Battelle Memorial Institute Photoacoustic spectroscopy sample array vessels and photoacoustic spectroscopy methods for using the same
WO2008117086A1 (fr) * 2007-03-23 2008-10-02 Attomarker Limited Procédé de fabrication de réseaux de biodétecteurs photoniques
CN113826007A (zh) * 2019-03-28 2021-12-21 传感器动力公司 使用湿-干生物分析技术通过剪切水平表面声波生物传感器检测心肌钙蛋白或生物标志物
WO2022038586A2 (fr) 2020-08-17 2022-02-24 Cor Sync Desenvolvimento De Sistemas Ltda Dispositif et procédé de mesure du niveau de biomarqueurs et de pathogènes dans des substances

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US8076161B2 (en) * 2007-05-31 2011-12-13 Canon Kabushiki Kaisha Target substance detection kit and target substance detection method
JP4806699B2 (ja) * 2008-08-01 2011-11-02 日本電信電話株式会社 マイクロチップ及び該マイクロチップを用いた抗体固定化方法

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US4900662A (en) * 1987-07-21 1990-02-13 International Immunoassay Laboratories, Inc. CK-MM myocardial infarction immunoassay
EP0384130A2 (fr) * 1987-07-21 1990-08-29 International Immunoassay Laboratories, Inc. Immunoessai pour la détection de l'infarctus du myocarde
WO1990005305A1 (fr) * 1988-11-10 1990-05-17 Pharmacia Ab Ensemble capteur a resonance de plasmon de surface et son utilisation dans des systemes de biocapteurs
WO1991001498A1 (fr) * 1989-07-21 1991-02-07 Vioclone Biologicals Inc. Emploi de la creatine-kinase et de son isozyme ck-mb ainsi que de la chaine legere 1 de myosine ventriculaire humaine dans le diagnostic de l'insuffisance cardiaque
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* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP1019717A4 (fr) * 1997-09-18 2004-09-01 Univ Utah Res Found Dispositif et procede de diagnostic
US6673562B2 (en) 2000-08-24 2004-01-06 Spectral Diagnostics, Inc. Differential immunoassay
WO2002023191A1 (fr) * 2000-09-12 2002-03-21 University Of Sydney Dosage diagnostique
WO2002057776A3 (fr) * 2001-01-19 2003-05-08 Quantech Ltd Systeme et procede pour reduire l'interference des particules lors de la detection d'analytes fluides
US6870626B2 (en) 2001-11-13 2005-03-22 Battelle Memorial Institute Array-based photoacoustic spectroscopy
WO2003042669A1 (fr) * 2001-11-13 2003-05-22 Battelle Memorial Institute Spectroscopie photo-acoustique effectuee avec un reseau
US6873415B2 (en) 2001-11-13 2005-03-29 Battelle Memorial Institute Photoacoustic spectroscopy sample array vessel and photoacoustic spectroscopy method for using the same
US6999174B2 (en) 2001-11-13 2006-02-14 Battelle Memorial Institute Photoacoustic spectroscopy sample array vessels and photoacoustic spectroscopy methods for using the same
WO2004038415A1 (fr) * 2002-10-24 2004-05-06 Biacore Ab Test avec des ligands co-immobilises
WO2008117086A1 (fr) * 2007-03-23 2008-10-02 Attomarker Limited Procédé de fabrication de réseaux de biodétecteurs photoniques
US8394648B2 (en) 2007-03-23 2013-03-12 Attomarker Limited Method of fabrication of photonic biosensor arrays
CN113826007A (zh) * 2019-03-28 2021-12-21 传感器动力公司 使用湿-干生物分析技术通过剪切水平表面声波生物传感器检测心肌钙蛋白或生物标志物
WO2022038586A2 (fr) 2020-08-17 2022-02-24 Cor Sync Desenvolvimento De Sistemas Ltda Dispositif et procédé de mesure du niveau de biomarqueurs et de pathogènes dans des substances
WO2022038586A3 (fr) * 2020-08-17 2022-04-14 Cor Sync Desenvolvimento De Sistemas Ltda Dispositif et procédé de mesure du niveau de biomarqueurs et de pathogènes dans des substances

Also Published As

Publication number Publication date
AU656202B2 (en) 1995-01-27
JPH06508204A (ja) 1994-09-14
SE9101735D0 (sv) 1991-06-07
AU1975692A (en) 1993-01-08
EP0588891A1 (fr) 1994-03-30
CA2110705A1 (fr) 1992-12-10

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