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鼠源性NAA10单克隆抗体C7,3A9,5E3的制备与初步鉴定

, PP. 1341-1345

Keywords: NAA10,单克隆抗体,制备,鉴定

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Abstract:

目的制备乙酰基转移酶NAA10单克隆抗体并进行初步鉴定。方法利用PCR扩增技术由结肠癌细胞株HT-29中克隆出NAA10全长cDNA。将上述cDNA与原核表达载体pGEX-4T1重组,并将重组体转化大肠杆菌BL-21(DE3),随后行融合蛋白GST-NAA10的表诱导达,用GlutathioneSepharose4B进行纯化。用纯化的GST-NAA10融合蛋白背部皮下多点注射免疫Balb/c小鼠,通过杂交瘤技术制备抗Naa10的单克隆抗体;随后利用ELISA鉴定其效价,Westernblotting鉴定特异性。结果本研究获得只与NAA10特异反应的单抗3株(C7、3A9和5E3),初步确定C7为IgM型,5E3为IgG2a型,3A9为IgM型;单抗C7、5E3针对NAA10具有较高特异性,而3A9特异性相对较差。结论稳定表达的NAA10单克隆抗体细胞株建立成功。

References

[1]  Joo-Won Jeong, Moon-Kyoung Bae, Mee-Young Ahn,et al.Regulation and destabilization of HIF-1αby ARD1-mediated acetylation[J].Cell,2002,111:709-720.
[2]  Polevoda B, Arnesen T, Sherman F.A synopsisof eukaryotic N-alpha-terminal acetyltransferases:nomenclature, subunits and substrate[J].BMC Proc,2009,3(Suppl 6):S2.
[3]  闵立,曾妍,寿成超. 乙酰转移酶基因NAA10的生物学功能及其在肿瘤中的作用[J].中国生物化学与分子生物学报,2013,29(10):902-909.
[4]  范璐,谭德勇.N-乙酰基转移酶ARD1基因及其与肿瘤关系研究进展[J].生物技术通讯,2013,24(4):558-561.
[5]  Polevoda B, Sherman F.N-terminal acetyltransferases and sequence requirements for N-terminal acetylation of eukaryotic protein[J]. J Mol Biol, 2003, 325(4): 595-622.
[6]  Meinnel T, Peynot P, Giglione C. Processed N termini of mature protein in higher eukaryotes and their major contribution to dynamic proteomics[J]. Biochimie, 2005, 87(8): 701-712.
[7]  Bilton R, Mazure N, Trottier E,et al.Arrest-defective-1 protein,an acetyltransferase,does not alter stability of hypoxia-inducible Factor(HIF)-1 and is not induced by hypoxia or HIF[J].J Biol Chem,2005,280(35),31132-31140.
[8]  Hua KT,Tan CT,Johansson G,et al.N-acetyltransferase 10 protein suppresses cancer metastasis by binding PIX proteins and inhibiting Cdc42/Rac1 activity[J].Cancer Cell,2011,19(2):218-231.
[9]  Kuo HP,Lee DF,Chen CT,et al.ARD1 stabilization of TSC2 suppresses tumorigenesis through the mTOR signaling pathway[J].Sci Signal, 2010,3(108):ra9.
[10]  余敏,王泽华,龚军丽,等.用于人乳腺癌病理检测的抗人hARD1单克隆抗体的制备[J].生物工程学报,2010,26(1):57-62.
[11]  白松,周媛媛,田风,等.ARD1在人大肠腺癌细胞SW620、LS-174T和HCT-8中的表达[J].南方医科大学学报,2011,31(5):914-916.

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