▎ 摘　　要

NOVELTY - Hypoxanthine/aminopterin/thymidine (HAT) semi-solid screening culture medium comprises 9-15 pts. wt. biological protein hydrolyzate, 5-9 pts. wt. carrier film, 6-15 pts. wt. fetal calf serum and 6-15 pts. wt. polyethylene glycol. USE - The HAT semi-solid screening culture medium is useful for screening monoclonal antibody and culturing hybridoma cell culture. DETAILED DESCRIPTION - INDEPENDENT CLAIMS are included for the following: (1) preparation method of HAT semi-solid screening culture medium for preparing monoclonal antibody, involves extracting the biological protein hydrolyzate, preparing carrier film and compounding the mixture; and (2) usage method of the monoclonal antibody composition for preparing HAT semi-solid screened the monoclonal antibody composition, involves returning 90-100 ml (for fusion of 1-2 mice) semi-solid medium to room temperature or equilibrating in a cell incubator, centrifuging the fused cells at 1500 revolutions per minute (rpm), removing polyethylene glycol (PEG) or fusion buffer for 10 minutes, resuspending the cells with 10 ml base medium, adding the cells to the semi-solid medium to mix (or gently mixing with a rotor for 5 minutes), placing in 3.5 cm dishes after the fused cells and the culture medium are fully mixed, culturing in a sterile incubator with a carbon dioxide content of 5% and a constant temperature of 37degreesC, where the negative control is set as SP2/0 well, observing SP2 on the next day, normally stopping growing SP2/0 for 4 days of culture, observing obvious cell colonies under the microscope, after 7 days of culture, where cell colonies are visible to the naked eye, selecting spots of cell colonies, where the selected spots are single cloning, subculturing the monoclonal hybridoma cell line in the order of 96-well-24-well-6-well-cell plate, performing enzyme-linked immunosorbent assay (ELISA) detection during the subculture process to retain the high-titer and stable hybridoma cell line, harvesting the culture supernatant after the hybridoma cell line is expanded and cultured, identifying the antibody activity after affinity purification, inoculating identified hybridoma cell line into the peritoneal cavity of paraffin immunosuppressed mice, inducing ascites on the 7th day, collecting the ascites, and separating the ascites by protein G affinity purification.