QUANTIFICATION OF THE SECRETORY GLYCOPROTEINS OF THE SUBCOMMISSURAL ORGAN BY A SENSITIVE SANDWICH ELISA WITH A POLYCLONAL ANTIBODY AND A SET OF MONOCLONAL-ANTIBODIES AGAINST THE BOVINE REISSNERS-FIBER

The subcommissural organ (SCO) is an ependymal brain gland that releas es glycoproteins into the ventricular cerebrospinal fluid where they c ondense to form the Reissner's fiber (RF). We have developed a highly sensitive and specific two-antibody sandwich enzyme-linked immunosorbe nt assay (ELISA) for the quantification of the bovine SCO secretory ma terial. The assay was based on the use of the IgG fraction of a polycl onal antiserum against the bovine RF as capture antibody and a pool of three peroxidase-labeled monoclonal antibodies that recognize non-ove rlapping epitopes of the RF glycoproteins as detection antibody. The d etection limit was 1 ng/ml and the working range extended from 1 to 40 00 ng/ml. The calibration curve, generated with RF glycoproteins, show ed two linear segments: one of low sensitivity, ranging from 1 to 125 ng/ml, and the other of high sensitivity between 125 and 4000 ng/ml. T his assay was highly reproducible (mean intra- and interassay coeffici ent of variation 2.2% and 5.3%, respectively) and its detectability an d sensitivity were higher than those of ELISAs using exclusively eithe r polyclonal or monoclonal antibodies against RF glycoproteins. The as say succeeded in detecting and measuring secretory material in crude e xtracts of bovine SCO, culture medium supernatant of SCO explants and incubation medium of bovine RF; however, soluble secretory material wa s not detected in bovine cerebrospinal fluid.