Specific monoclonal antibodies of coding regions and constant regions of goose immunoglobulin, and applications thereof

[0100] Example 1 Cloning of goose immunoglobulin λ chain, α chain, μ chain, υ chain and its constant region cDNA sequence

[0101] 1. Extraction of total RNA from goose spleen

[0102] Fresh goose spleen was taken, immediately ground in liquid nitrogen, and RNA was extracted using Trizol reagent (Invitrogen). The specific operation is as follows: add 1ml Trizol reagent per 100mg tissue, and place at room temperature for 10min; add 0.2ml chloroform for every 1ml TRIzol used, shake vigorously for 15 seconds, place at room temperature for 15min, and centrifuge at 12000rpm for 15min at 4°C; transfer the aqueous phase to a new tube, and use Precipitate the RNA in the aqueous phase with 2.5 times the volume of isopropanol at -20°C, then centrifuge at 12,000 rpm for 15 minutes at 4°C; wash the RNA pellet with 1ml of 75% ethanol (add at least 1ml of 75% ethanol for every 1ml of TRIzol used), at 4°C Centrifuge at 10,000 rpm for 5 minutes, discard the supernatant; place at room temper...

[0143] Embodiment 2 Preparation of anti-goose immunoglobulin constant region monoclonal antibody

[0144] 1. Preparation of immunogen

[0145] The specific operation of protein A affinity chromatography to purify goose serum immunoglobulin is as follows: take out the column filled with Protein A filler (GenScript, Nanjing), drain the liquid, and wash with deionized water for 2 to 3 times (the liquid added to the column is Filtered through a 0.45μm filter), and washed with Wash Buffer (0.15M NaCl, 20mM Na 2 HPO 4 , pH 8.0) equilibrate the column for 2-3 times; add serum diluted with 2 times of Wash Buffer, rotate the column continuously to fully combine the target protein with the filler, and collect the effluent; add 1ml of Wash Buffer, wash 7-8 times, and collect the effluent Add 1ml of Elution Buffer (citric acid 0.1M, pH 2.8), elute 7 to 8 times, collect the effluent, and immediately neutralize it to pH 7.4 with 1M Tris-base solution; SDS-PAGE detects the protein purifica...

[0159] Monoclonal antibody detection specific antibody after embodiment 4 enzyme labeling

[0160] HRP-labeled anti-goose immunoglobulin lambda chain constant region monoclonal antibody (secreted by hybridoma cell line 1B11) detection specific antibody:

[0161] Take 0.1mg BSA as immunogen to immunize adult geese by intramuscular injection, collect blood once every 4 days, separate serum, use conventional indirect ELISA method to detect specific antibodies produced in goose body, use BSA as antigen to coat microtiter plate, 5 % skim milk was used as blocking solution, 1:10 goose serum was used as primary antibody, and HRP-labeled 1B11 monoclonal antibody was used as secondary antibody to determine the growth and decline of specific antibodies in the body of goose immunized with BSA ( Figure 16 a). Similarly, the indirect ELISA method was used to measure goose serum immunized with gosling plague virus (GPV) virus-like particles (VLP) preserved in our laboratory, and the GPV v...