GEL FIBERGLASS AS A NEW MATRIX OF AFFINITY-CHROMATOGRAPHY COLUMNS FORISOLATION OF COLON CANCER-ASSOCIATED ANTIGENS

Gel fiberglass (GFG), a new affinity biosensor, was used to isolate hu man p53 antigen with rabbit anti-rat p53 IgG. The biosensor was prepar ed as a membrane from glass fibers covered with oxysilanes. A thin lay er of protein, trapped in gel glass during its preparation, is deposit ed on the surface of a lattice of glass fibers. In such conditions, a maximum number of protein molecules may contact external agents percol ated through a membrane. The membranes demonstrate high stability and can be stored in dry conditions or several months at room temperature. Columns for affinity chromatography were prepared from the GFG membra nes and were used to isolate various proteins, including the tumor-ass ociated antigens (TAA). The capacity of such columns was calculated as the amount mg of protein isolated from 1 ml of TAA-containing serum. In colon cancer patients, up to 5-6 mg TAA were extracted from 1 mi of sera. Two main components of cytoplasmic TAA isolated in our experime nts were p64 and p53 proteins. Their concentration was determined by H PLC. The p53 protein has been isolated from the serum of cancer patien ts in the highest concentration yet reported, up to 3-4 mg/ml. In our previous studies, isolation of p53 protein was based on its affinity r eaction with anti-p53 IgG generated against antigens of the same speci es. Herein, we report for the first time the capability to isolate hum an p53 antigen using GFG columns with entrapped anti-rat p53 IgG. Bloo d levels of p53 antigen isolated were very similar in both experiments . This has both theoretical and practical significance, demonstrating that the GFG membranes have great potential for isolating macromolecul es utilizing various ligands. The finding facilitates an easy and high ly effective method to isolate antigens from different organs, both an imal and human, which can be used for important goals including diagno sis, therapy and generation of specific antibodies.