HAT cell culture medium is a chemically defined selective medium specifically designed for hybridoma cell selection. Its core components achieve highly efficient and specific screening of fusion cells through the synergistic action of hypoxanthine (H), aminopterin (A), and thymidine (T). This medium employs aminopterin to block primary DNA synthesis pathways while relying on the hypoxanthine-thymidine nucleoside salvage pathway. This ensures survival and proliferation of only hybridoma cells possessing HGPRT enzyme activity, while unfused myeloma cells and B lymphocytes are selectively eliminated due to metabolic defects.