Method of evaluation of the relative risk of developing atherosclerosis in patients

Abstract

The present invention relates to a method for determining the amount of circulating CD36 protein or a fraction thereof which is present in cell-free plasma, preferably in a high molecular weight plasma fraction, such as a lipoprotein fraction selected from Low Density Lipoprotein, Intermediate Density Lipoprotein, and Very Low Density Lipoprotein using an immunological method which comprises the steps of (i) providing a plasma sample to be investigated, (ii) providing an anti-CD36 antibody, (iii) exposing the sample to be investigated to the antibody, and (iv) detecting and quantifying the amount of CD36 which binds to the antibody.

1 - 15 . (canceled) 
     
     
         16 . A solid phase enzyme immunoassay, comprising
 (a) a cell-free plasma sample applied to the solid phase comprising a high molecular weight lipoprotein fraction from a patient at risk of developing a disease state selected from atherosclerosis, atherothrombosis, microangiopathy, metabolic syndrome, non-alcoholic steatohepatitis, insulin resistance, diabetes, and combinations thereof, wherein high molecular weight (lipid-protein) complexes in the fraction are degraded; and   (b) a CD36-antibody complex comprising (i) an anti-human CD36 antibody which is bound to the solid phase and (ii) a CD36 protein complex from the high molecular weight lipoprotein fraction.   
     
     
         17 . The solid phase enzyme immunoassay of  claim 16 , comprising
 (c) a labelled compound having specific binding affinity for said complex and which is bound to said complex, optionally wherein the label is optionally selected from a radioactive label, a chemiluminescent label, a fluorescent dye, and an enzyme label.   
     
     
         18 . The solid phase enzyme immunoassay of  claim 17 , wherein the labelled compound of (c) is a secondary antibody. 
     
     
         19 . The solid phase enzyme immunoassay of  claim 16 , wherein the CD36 of (b) is bound to Low Density Lipoprotein, Intermediate Density Lipoprotein, or Very Low Density Lipoprotein which is present in the high molecular weight fraction. 
     
     
         20 . The solid phase enzyme immunoassay of  claim 16 , wherein the molecular weight of said high molecular weight complexes is within 440,000-2,000,000 g/mol. 
     
     
         21 . The solid phase enzyme immunoassay of  claim 20 , wherein the molecular weight of said high molecular weight complexes is around 1,000,000 g/mol. 
     
     
         22 . The solid phase enzyme immunoassay of  claim 16 , wherein the patient is selected from the group consisting of DM2 patients, obese DM2 patients, and healthy relatives of DM 2  patients and non-diabetic obese persons. 
     
     
         23 . The solid phase enzyme immunoassay of  claim 16 , wherein CD36 levels in the patient sample are increased by at least 250% relative to a reference level of circulating CD36 in healthy subjects. 
     
     
         24 . The solid phase enzyme immunoassay of  claim 16 , wherein the solid phase is a microtiter plate. 
     
     
         25 . The solid phase enzyme immunoassay of  claim 16 , wherein the CD36 protein is a CD36-lipoprotein complex. 
     
     
         26 . A method of measuring a circulating CD36 level in a patient, comprising measuring the circulating CD36 level in a solid phase enzyme immunoassay according to  claim 16 .