IMMUNOBLOT ANALYSIS OF URINARY FIBRONECTIN OF NORMAL AND PASSIVE HEYMANN NEPHRITIS RATS

Immunoblot analysis for urinary fibronectin (FN) of normal and passive Heymann nephritis (PHN) rats was performed. When 2-5% skim milk or 2- 10% bovine serum albumin solution was used as a blocking buffer for th e immunoblot analysis, remarkable nonspecific interactions were observ ed between many urinary proteins and immunoglobulins used as a primary as well as a secondary antibody. Blocking of the trnasblot membrane w ith a 2% gamma-globulin solution prevented such nonspecific interactio ns, As a result, two FN fragments, not intact FN, with molecular masse s of 55- and 65-kDa were detected in all the normal rat urine samples. In PHN rat urine, intact FN and several FN fragments of 100-200-kDa w ere evident instead of the 55- and 65-kDa FN fragments. These results indicate that metabolism of FN and excretion of the FN metabolites int o urine may be changed during the development of the renal disease.