The invention discloses a preparation method of an immunoaffinity column. The immunoaffinity column is a compound affinity column which is mainly prepared from three types of immunoaffinity columns through mixed packing in a volume ratio being 1:1:1, and the three types of immunoaffinity columns are prepared from three antibodies and a coupling matrix through coupling and wet packing. The column capacity of the immunoaffinity column is 2,500 ng, the immunoaffinity column is good in purification effect and can be repeatedly used three times, and average recovery rate is 67%-107%. The compound immunoaffinity column has good specific adsorption property for at least 12 sulfonamides including sulfaquinoxaline, sulfadiazine, sulfapyridine, sulfametoxydiazine, sulfadimidine, sulfamethoxazole, sulfafurazole, sulfadimethoxypyrimidine, sulfathiazole, sulfamerazine, sodium N-sulphanilamidate and sulfamonomethoxine. The immunoaffinity column which is high in coupling rate, specific adsorption property, column capacity and recovery rate is prepared, and a sensitive, rapid and simple analysis method capable of detecting residues of the 12 sulfonamides simultaneously is provided.