A method for expressing and preparing a bivalent bispecific antibody, wherein said antibody comprises: a light chain and a heavy chain of the first antibody that specifically binds to a first antigen; a light chain and the VH+CH1 portion of a heavy chain of the second antibody that specifically binds to a second antigen, wherein the C-terminus of the CH1 domain is fused to an Intein-N moiety (IN); an Fc chain from the second antibody, wherein the N-terminus of the Fc chain is linked to an Intein-C moiety (IC); a knob and a corresponding hole capable of associating with each other, the knob and hole being located in the interface of the CH3 domain of the heavy chain of the first antibody and in the interface of the CH3 domain of the Fc chain or vice versa; wherein the method comprises the following steps: A1: obtaining and amplifying "portion A antibody", which comprises the light chain and the heavy chain of the first antibody, and the Fc chain of the heavy chain of the second antibody having IC fused to the N terminus; A2: constructing one or more mammalian cell expression vectors by whole-gene synthesis, wherein the vector or vectors comprises the coding sequences amplified in step A1; A3: obtaining the portion A antibody by expressing the vector or vectors prepared in step A2 in stably or transiently transfected mammalian cells; B1: obtaining and amplifying the coding sequences of "portion B antibody", which comprises the light chain and VH+CH1 domains of the second antibody, with the IN moiety linked to the CH1 C-terminus; B2: constructing one or more mammalian cell expression vector by whole-gene synthesis, wherein the vector or vectors comprises the coding sequences amplified in step B1; B3: obtaining the portion B antibody by expressing the vector or vectors prepared in step B2 in stably or transiently transfected mammalian cells; and C: obtaining the bivalent bispecific antibody by subjecting the portion A antibody obtained in step A3 and the portion B antibody obtained in step B3 to trans-splicing in vitro. a light chain and a heavy chain of the first antibody that specifically binds to a first antigen; a light chain and the VH+CH1 portion of a heavy chain of the second antibody that specifically binds to a second antigen, wherein the C-terminus of the CH1 domain is fused to an Intein-N moiety (IN); an Fc chain from the second antibody, wherein the N-terminus of the Fc chain is linked to an Intein-C moiety (IC); a knob and a corresponding hole capable of associating with each other, the knob and hole being located in the interface of the CH3 domain of the heavy chain of the first antibody and in the interface of the CH3 domain of the Fc chain or vice versa; A1: obtaining and amplifying "portion A antibody", which comprises the light chain and the heavy chain of the first antibody, and the Fc chain of the heavy chain of the second antibody having IC fused to the N terminus; A2: constructing one or more mammalian cell expression vectors by whole-gene synthesis, wherein the vector or vectors comprises the coding sequences amplified in step A1; A3: obtaining the portion A antibody by expressing the vector or vectors prepared in step A2 in stably or transiently transfected mammalian cells; B1: obtaining and amplifying the coding sequences of "portion B antibody", which comprises the light chain and VH+CH1 domains of the second antibody, with the IN moiety linked to the CH1 C-terminus; B2: constructing one or more mammalian cell expression vector by whole-gene synthesis, wherein the vector or vectors comprises the coding sequences amplified in step B1; B3: obtaining the portion B antibody by expressing the vector or vectors prepared in step B2 in stably or transiently transfected mammalian cells; and C: obtaining the bivalent bispecific antibody by subjecting the portion A antibody obtained in step A3 and the portion B antibody obtained in step B3 to trans-splicing in vitro. <EOS>