De-fucosylated fully humanized monoclonal antibody and applications thereof

[0045] Example 1 Preparation, expression, purification and detection of antibodies

[0046] The preparation, expression, purification of Fab and IgG1 antibody, and the method of converting Fab into IgG1 were carried out according to the method reported in the literature (3);

[0047] The m336-gH gene line was obtained by mutating the 59th amino acid of the heavy chain variable region of m336 from S to N through a site-directed mutagenesis kit, and was prepared using expression and purification conditions similar to those of m336;

[0048] The specific method of Fab preparation is as follows: transfer the plasmid containing the clone m336 or m336-gH into HB2151 competent cells, pick a single colony from the ampicillin plate grown overnight, inoculate the SB bacterial culture medium, and express under the induction condition of 30 degrees IPTG 10-12 hours; Bacteria are harvested and Fabs are purified from them. IgG1 antibody was prepared and expressed by stably transfected and ...