A kind of monoclonal antibody of ultraviolet excision repair protein and its application
A monoclonal antibody, repair protein technology, applied in anti-animal/human immunoglobulin, biochemical equipment and methods, material testing products, etc., can solve problems such as differences
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Embodiment 1
[0020] Example 1: Obtaining the hybridoma cell line 5H1-A10-A7 and the monoclonal antibody it produces.
[0021] (1), animal immunity
[0022] 6-8 week old female BALB / c mice were immunized with recombinant human hHR23b protein as antigen, and three immunization injections were performed according to the pre-established immunization method.
[0023] The sequence structure of the antigenic protein is:
[0024] MQVTLKTLQQQTFKIDIDPEETVKALKEKIESEKGKDAFPVAGQKLIYAGKILNDDTALKEYKIDEKNFVVVMVTKPKAVSTPAPATTQQSAPASTTAVTSSTTTTVAQAPTPVPALAPTSTPASITPASATASSEPAPASAAKQEKPAEKPAETPVATSPTATDSTSGDSSRSNLFEDATSALVTGQSYENMVTEIMSMGYEREQVIAALRASFNNPDRAVEYLLMGIPGDRESQAVVDPPQAASTGAPQSSAVAAAAATTTATTTTTSSGGHPLEFLRNQPQFQQMRQIIQQNPSLLPALLQQIGRENPQLLQQISQHQEHFIQMLNEPVQEAGGQGGGGGGGSGGIAEAGSGHMNYIQVTPQEKEAIERLKALGFPEGLVIQAYFACEKNENLAANFLLQQNFDED
[0025] For the first immunization, 0.2ml emulsion (100ul purified recombinant protein hHR23b (diluted with 1×PBS) + 100ul complete Freund's adjuvant, containing 100u...
Embodiment 2
[0037] Example 2: Identification and Application of Monoclonal Antibody
[0038] 1. Western blot detection and identification of mouse anti-hHR23b monoclonal antibody (anti-hHR23b)
[0039] Extract the cell lysates of various tumor cell lines, adjust the total protein concentration to 2mg / ml, load the samples into 12% SDS-PAGE gel holes, 20ug / well, and transfer the proteins in the gel to PVDF membrane after electrophoresis , after blocking with 5% skimmed milk powder, immunostaining was carried out with the anti-hHR23b antibody prepared in Example 1: add the anti-hHR23b prepared and purified in step (3) of Example 1, the working concentration was 1:1000, and incubated overnight at 4 degrees , and then add HRP-labeled goat anti-mouse antibody at a working concentration of 1:20000, and react at 37°C for 1 hr, and a 55Kd band appears in each lysate (see figure 2 ), consistent with literature reports, proving that this antibody is a specific antibody against hHR23b; the antibody...
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