Process for analyzing annexin-V in urine, and application thereof

Abstract

The present invention provides a process and regent for analyzing annexin-V, wherein the measurement of a concentration of annexin-V can be easily carried out without need for addition of chemicals for inhibiting the bonding of various proteins with calcium ion and for adjusting a specimen solution to the specimen at a measuring stage, and a process and medicine for diagnosing an internal organ disorder based on the analyzing process and regent. A urine is brought into contact with an anti-annexin-V monoclonal antibody to perform an antigen-antibody reaction of annexin-V in the urine with the anti-annexin-V monoclonal antibody, thereby forming an annexin-V antigen/anti-annexin-V monoclonal antibody complex, and the amount of the formed annexin-V antigen/anti-annexin-V monoclonal antibody complex is quantitatively measured. Thus, it is possible to carry out the measurement of the concentration of annexin-V in the urine, and it is possible to carry out the diagnosis of an internal organ disorder such as a disseminated intravascular coagulation syndrome, the diagnose of acute nephritis and the like by a analyzed value of concentration of annexin-V measured in the urine.

What is claimed is  
     
         1 . A process for analyzing annexin-V in a urine, comprising the steps of bringing a urine into contact with an anti-annexin-V monoclonal antibody to perform an antigen-antibody reaction of annexin-V being present in the urine with said anti-annexin-V monoclonal antibody, thereby forming an annexin-V antigen/anti-annexin-V monoclonal antibody complex; and quantitatively measuring the amount of said formed annexin-V antigen/anti-annexin-V monoclonal antibody complex.  
     
     
         2 . A process for analyzing annexin-V in a urine according to  claim 1 , wherein said anti-annexin-V monoclonal antibody is produced by a hybridoma cell line deposited under the number of FERM BP-5284 or a hybridoma cell line deposited under the number of FERM BP-5286 in the International Depositary Authority for the deposit of microorganism.  
     
     
         3 . A process for analyzing annexin-V in a urine, comprising the steps of bringing a urine into contact with a first anti-annexin-V monoclonal antibody fixed in a solid phase to perform an antigen-antibody reaction of annexin-V being present in the urine with said first anti-annexin-V monoclonal antibody, thereby forming an annexin-V antigen/first anti-annexin-V monoclonal antibody complex; bonding an anti-dog-annexin-V polyclonal labeled antibody or a second anti-annexin-V monoclonal labeled antibody to the annexin-V antigen of said formed antibody complex to form a bonded antibody complex/anti-annexin-V polyclonal or second anti-annexin-V monoclonal labeled antibody product; and quantitatively measuring the amount of said formed labeled antibody bonded product.  
     
     
         4 . A process for analyzing annexin-V in a urine according to  claim 3 , wherein said first anti-annexin-V monoclonal antibody is produced by a hybridoma cell line of reception No.FERM BP-5284 or reception No.FERM BP-5286 in International Microorganism Consignment Authority, and said second anti-annexin-V monoclonal labeled antibody is formed by a product produced by the hybridoma cell line deposited under the number of FERM BP-5284 or FERM BP-5286 in the International Depositary Authority for the deposit of microorganism, said first anti-annexin-V monoclonal antibody and the second anti-annexin-V monoclonal antibody forming said second anti-annexin-V monoclonal labeled antibody being antibodies different from each other.  
     
     
         5 . A regent for analyzing annexin-V in a urine, comprising a first anti-human-annexin-V monoclonal antibody having a specificity to an antigen determinant site on the protein of a human annexin-V antigen fixed in a solid phase, and a second anti-human-annexin-V monoclonal labeled antibody or an anti-dog-annexin-V polyclonal labeled antibody having a binding specificity to the antigen determinant site on the protein of the human annexin-V antigen.  
     
     
         6 . A regent for analyzing annexin-V in a urine according to  claim 5 , wherein said first anti-human-annexin-V monoclonal antibody is produced by a hybridoma cell line deposited under the number FERM BP-5284 or FERM BP-5286 in the International Depositary Authority for the deposit of microorganism, and said second anti-annexin-V monoclonal labeled antibody is formed by a product produced by the hybridoma cell line deposited under the number FERM BP-5284 or FERM BP-5286 in the International Depositary Authority for the deposit of microorganism, said first anti-annexin-V monoclonal antibody and the second anti-annexin-V monoclonal antibody forming said second anti-annexin-V monoclonal labeled antibody being antibodies different from each other.  
     
     
         7 . A process for diagnosing an organopathy, comprising the steps of measuring a concentration of annexin-V in a urine, and comparing the measured value of the concentration of the annexin-V in the urine with a normal value of concentration of annexin-V.  
     
     
         8 . A process for diagnosing an organopathy according to  claim 7 , wherein the urine is brought into contact with an anti-annexin-V monoclonal antibody to perform an antigen-antibody reaction of annexin-V in said urine with said anti-annexin-V monoclonal antibody, thereby forming an annexin-V antigen/anti-annexin-V monoclonal antibody complex, and quantitatively measuring the amount of said formed annexin-V antigen/anti-annexin-V monoclonal antibody complex.  
     
     
         9 . A process for diagnosing an organopathy according to  claim 8 , wherein said anti-annexin-V monoclonal antibody is produced by a hybridoma cell line deposited under the number FERM BP-5284 or FERM BP-5286 in the International Depositary Authority for the deposit of microorganism.  
     
     
         10 . A process for diagnosing an organopathy by use of a urine, comprising the steps of bringing a urine into contact with a first anti-annexin-V monoclonal antibody to perform an antigen/antibody reaction of annexin-V in the urine with said first anti-annexin-V monoclonal antibody, thereby forming an annexin-V antibody/first anti-annexin-V monoclonal antibody complex; bonding an anti-dog-annexin-V polyclonal labeled antibody or a second anti-annexin-V monoclonal labeled antibody to the annexin-V antigen of said formed antibody complex to form a bonded product of anti-dog-annexin-V polyclonal labeled antibody or second anti-annexin-V monoclonal labeled antibody/annexin-V antigen/antibody complex; and quantitatively measuring the amount of said formed bonded product of anti-dog-annexin-V polyclonal labeled antibody or second anti-annexin-V monoclonal labeled antibody/annexin-V antigen/antibody complex.  
     
     
         11 . A process for diagnosing an organopathy by use of a urine according to  claim 10 , wherein said first anti-annexin-V monoclonal antibody is produced by a hybridoma cell line deposited under the number FERM BP-5284 or FERM BP-5286 in the International Depositary Authority for the deposit of microorganism, and said second anti-annexin-V monoclonal labeled antibody is formed by a product produced by the hybridoma cell line deposited under the number FERM BP-5284 or FERM BP-5286 in the International Depositary Authority for the deposit of microorganism, said first anti-annexin-V monoclonal antibody and the second anti-annexin-V monoclonal antibody forming said second anti-annexin-V monoclonal labeled antibody being antibodies different from each other.  
     
     
         12 . A process for diagnosing an organopathy according to any of  claims 7  to  11 , wherein the internal disorder is accompanied by a disseminated intravascular coagulation syndrome.  
     
     
         13 . A medicine for diagnosing an organopathy using a urine, comprising a first anti-human-annexin-V monoclonal antibody having a binding specificity to an antigen determinant site on the protein of a human annexin-V antigen fixed in a solid phase, and a second anti-human-annexin-V monoclonal labeled antibody or an anti-dog-annexin-V polyclonal labeled antibody having a binding specificity to the antigen determinant site on the protein of the human annexin-V antigen.  
     
     
         14 . A medicine for diagnosing an organopathy using a urine according to  claim 13 , wherein said first anti-annexin-V monoclonal antibody is produced by a hybridoma cell line deposited under the number FERM BP-5284 or FERM BP-5286 in the International Depositary Authority for the deposit of microorganism, and said second anti-annexin-V monoclonal labeled antibody is formed by a product produced by the hybridoma cell line deposited under the number FERM BP-5284 or FERM BP-5286 in the International Depositary Authority for the deposit of microorganism, said first anti-annexin-V monoclonal antibody and the second anti-annexin-V monoclonal antibody forming said second anti-annexin-V monoclonal labeled antibody being antibodies different from each other.  
     
     
         15 . A medicine for diagnosing an organopathy using a urine according to  claim 13  or  14 , wherein the internal disorder is accompanied by a disseminated intravascular coagulation syndrome.  
     
     
         16 . A process for diagnosing acute nephritis by use of a urine, comprising the steps of measuring concentrations of annexin-V in samples of urine excreted at two different time points; determining a difference between measured values of concentrations of annexin-V in the samples of urine at said two different time points and measuring concentrations of annexin-V in samples of blood at said two different time points; determining a difference between measured values of concentrations of annexin-V in the samples of blood at said two different time points; and comparing said difference between measured values of concentrations of annexin-V in the samples of urine with said difference between measured values of concentrations of annexin-V in the blood.  
     
     
         17 . A process for diagnosing acute nephritis by use of a urine according to  claim 16 , wherein the concentration of annexin-V in the urine is measured by subjecting annexin-V of the antigen protein in the urine to an antigen-antibody reaction with a first anti-annexin-V monoclonal antibody to form an antibody complex, bonding a labeled anti-annexin-V polyclonal antibody or a second labeled anti-annexin-V monoclonal antibody to the annexin-V antigen of said formed antibody complex to form a bonded product of annexin-V antigen/antibody complex/anti-dog-annexin-V polyclonal antibody or second anti-annexin-V monoclonal antibody, and quantitatively measuring the amount of said formed antibody-coupled product.  
     
     
         18 . A process for diagnosing acute nephritis by use of a urine according to  claim 17 , wherein said first anti-annexin-V monoclonal antibody is produced by a hybridoma cell line deposited under the number FERM BP-5284 or FERM BP-5286 in the International Depositary Authority for the deposit of microorganism, and said second anti-annexin-V monoclonal labeled antibody is formed by a product produced by the hybridoma cell line deposited under the number FERM BP-5284 or FERM BP-5286 in the International Depositary Authority for the deposit of microorganism, said first anti-annexin-V monoclonal antibody and the second anti-annexin-V monoclonal antibody forming said second anti-annexin-V monoclonal labeled antibody being antibodies different from each other.  
     
     
         19 . A process for diagnosing acute nephritis by use of a urine, comprising the steps of bringing samples of urine taken at two different time points into contact with an anti-annexin-V monoclonal antibody to perform an antigen/antibody reaction of annexin-V in each of the urine samples with said anti-annexin-V monoclonal antibody, thereby forming an annexin-V antigen/anti-annexin-V monoclonal antibody complex; quantitatively measuring the amount of said formed annexin-V antigen/anti-annexin-V monoclonal antibody complex, thereby determining a concentration of annexin-V in each of the urine samples to determine a difference between the concentrations of annexin-V in the urine samples taken at said two different time points from the values of the concentrations of annexin-V in the urine samples taken at said two different time points; and on the other hand, bringing annexin-V present in a specimen derived from samples of blood drawn at said two different time points into an antigen/antibody reaction with an anti-annexin-V monoclonal antibody to form an annexin-V antigen/anti-annexin-V monoclonal antibody complex; quantitatively measuring the amount of said formed annexin-V antigen/anti-annexin-V monoclonal antibody complex, thereby determining a concentration of annexin-V in each of the blood samples to determine a difference between the concentrations of annexin-V in the blood samples drawn at said two different time points; and comparing the differences between the annexin-V concentrations in the urine samples and the blood samples taken at said two different time points with each other.  
     
     
         20 . A process for diagnosing acute nephritis by use of a urine according to  claim 19 , wherein said anti-annexin-V monoclonal antibody is produced by a hybridoma cell line deposited under the number FERM BP-5284 or a hybridoma cell line deposited under the number FERM BP-5286 in the International Depositary Authority for deposit of microorganism.  
     
     
         21 . A medicine for diagnosing acute nephritis by use of a urine, comprising a regent for analyzing annexin-V present in a urine, and a regent for analyzing annexin-V present in blood.  
     
     
         22 . A medicine for diagnosing acute nephritis by use of a urine according to  claim 21 , wherein said regent for analyzing annexin-V being present in the urine contains a first anti-annexin-V monoclonal fixed in a solid phase, and a second anti-annexin-V monoclonal labeled antibody or anti-annexin-V polyclonal labeled antibody, and said regent for analyzing annexin-V present in blood contains a third anti-annexin-V monoclonal fixed in a solid phase, a fourth anti-annexin-V monoclonal labeled antibody or anti-annexin-V polyclonal labeled antibody, and EDTA.  
     
     
         23 . A medicine for diagnosing acute nephritis by use of a urine according to  claim 22 , wherein in said regent for analyzing annexin-V being present in the urine, said first anti-annexin-V monoclonal is produced by a hybridoma cell line deposited under the number FERM BP-5284 or FERM BP-5286 in the International Depositary Authority for the deposit of microorganism, and said second anti-annexin-V monoclonal labeled antibody is formed by a product produced by the hybridoma cell line deposited under the number FERM BP-5284 or FERM BP-5286 in the International Depositary Authority for the deposit of microorganism, said first anti-annexin-V monoclonal antibody and said second anti-annexin-V monoclonal antibody being antibodies different from each other; and in said regent for analyzing annexin-V present in blood, said third anti-annexin-V monoclonal is produced by a hybridoma cell line deposited under the number FERM BP-5284 or FERM BP-5286 in the International Depositary Authority for the deposit of microorganism, and said fourth anti-annexin-V monoclonal labeled antibody is formed by a product produced by the hybridoma cell line deposited under the number FERM BP-5284 or FERM BP-5286 in the International Depositary Authority for the deposit of microorganism, said third anti-annexin-V monoclonal antibody and said fourth anti-annexin-V monoclonal antibody being antibodies different from each other.