[1]李星辰,李俊,包红朵,等.沙门氏菌噬菌体多克隆抗体制备及其检测方法[J].江苏农业学报,2025,(06):1233-1239.[doi:doi:10.3969/j.issn.1000-4440.2025.06.020]
　LI Xingchen,LI Jun,BAO Hongduo,et al.Preparation of polyclonal antibodies against Salmonella phage and establishment of their detection methods[J].,2025,(06):1233-1239.[doi:doi:10.3969/j.issn.1000-4440.2025.06.020]
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沙门氏菌噬菌体多克隆抗体制备及其检测方法()
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江苏农业学报[ISSN:1006-6977/CN:61-1281/TN]

卷:
期数:
2025年06期
页码:
1233-1239
栏目:
加工贮藏·质量安全
出版日期:
2025-06-30

文章信息/Info

Title:
Preparation of polyclonal antibodies against Salmonella phage and establishment of their detection methods
作者:
李星辰1; 2; 李俊2; 包红朵2; 何涛2; 王冉1; 2
(1.南京农业大学动物科技学院,江苏南京210095；2.江苏省农业科学院农产品质量安全与营养研究所,江苏南京210014)
Author(s):
LI Xingchen1; 2; LI Jun2; BAO Hongduo2; HE Tao2; WANG Ran1; 2
(1.College of Animal Science & Technology, Nanjing Agricultural University, Nanjing 210095, China；2.Institute of Food Safety and Nutrition, Jiangsu Academy of Agricultural Sciences, Nanjing 210014, China)
关键词:
沙门氏菌; 噬菌体; 抗体; 间接ELISA
Keywords:
Salmonella; phage; antibody; indirect ELISA
分类号:
R378.2+2；Q939.48
DOI:
doi:10.3969/j.issn.1000-4440.2025.06.020
文献标志码:
A
摘要:
噬菌体主要由蛋白质和核酸组成,进入动物体后可能会被机体认为是抗原,进而激发机体产生抗体。为深入研究噬菌体进入动物体内后抗体产生情况,本研究采用纯化后的广谱裂解性沙门氏菌噬菌体SCBS作为抗原,用新西兰大白兔开展多克隆抗体制备,最后成功获得了该噬菌体的多克隆抗体,并建立了SCBS噬菌体血清抗体的间接ELISA检测方法。抗原包被含量为1×109PFU/mL,用2%牛血清白蛋白(BSA)封闭3 h,血清稀释度1∶500,血清反应时间60 min,酶标二抗稀释至1∶12 000后作用45 min,TMB显色液反应15 min后测定吸光度(OD450)。该检测方法特异性良好,阳性临界值为0.372,阴性临界值为0.310；批内变异系数为2.30%～7.11%,批间变系数为1.37%～9.25%。
Abstract:
Phages are mainly composed of proteins and nucleic acids. When a bacteriophage enters an animal’s body, it may be recognized as an antigen, thereby triggering the production of antibodies. In order to further study the production of antibodies in the animal body after oral administration of phages, purified broad-spectrum lytic Salmonella phage SCBS was used as the antigen to challenge the New Zealand Great White Rabbit. The polyclonal antibodies against the phage SCBS were prepared, and the indirect ELISA method was established. The phage antibodies were detected as follows: phages were coated at a concentration of 1×109 PFU/mL, blocked with 2% bovine serum albumin (BSA) for 3 h, the hyperimmune serum (1∶500) was incubated for 60 min, then enzyme-labeled secondary antibody (1∶12 000) was incubated for 45 min, the absorbance (OD450) was measured after the substrate solution containing TMB reacted for 15 min. The established ELISA method demonstrated good specificity. The positive and negative threshold values were 0.372 and 0.310. The intra-assay coefficient of variation ranged from 2.30% to 7.11%, and the inter-assay coefficient of variation ranged from 1.37% to 9.25%.

参考文献/References:

[1]SRIVASTAVA A S, KAIDO T, CARRIER E. Immunological factors that affect the in vivo fate of T7 phage in the mouse[J]. Journal of Virological Methods,2004,115(1):99-104.
[2]WANG J Y, MENG W X, ZHANG K C, et al. Topically applied bacteriophage to control multi-drug resistant Pseudomonas aeruginosa-infected wounds in a New Zealand rabbit model[J]. Frontiers in Microbiology,2022,13:1031101.
[3]AHMED A M, SHIMAMOTO T. Isolation and molecular characterization of Salmonella enterica,Escherichia coli O157:H7 and Shigella spp. from meat and dairy products in Egypt[J]. International Journal of Food Microbiology,2014,168:57-62.
[4]AMARILLAS L, RUB-RANGEL L, CHAIDEZ C, et al. Isolation and characterization of phiLLS,a novel phage with potential biocontrol agent against multidrug-resistant Escherichia coli[J]. Frontiers in Microbiology,2017,8:1355.
[5]李胜,李广辉,张晓璐,等. 2020-2021年贵德县喜马拉雅旱獭血清中特异性鼠疫噬菌体抗体的检测[J]. 青海畜牧兽医杂志,2023,53(4):12-14,72.
[6]HODYRA-STEFANIAK K, MIERNIKIEWICZ P, DRAPAA J, et al. Mammalian host-versus-phage immune response determines phage fate in vivo[J]. Scientific Reports,2015,5:14802.
[7]JON＇CZYK-MATYSIAK E, USIAK-SZELACHOWSKA M, KAK M, et al. The effect of bacteriophage preparations on intracellular killing of bacteria by phagocytes[J]. Journal of Immunology Research,2015,2015:482863.
[8]Z·ACZEK M, USIAK-SZELACHOWSKA M, JON＇CZYK-MATYSIAK E, et al. Antibody production in response to staphylococcal MS-1 phage cocktail in patients undergoing phage therapy[J]. Frontiers in Microbiology,2016,7:1681.
[9]GAN S D, PATEL K R. Enzyme immunoassay and enzyme-linked immunosorbent assay[J]. Journal of Investigative Dermatology,2013,133(9):1-3.
[10]JOHNSON R P, GYLES C L, HUFF W E, et al. Bacteriophages for prophylaxis and therapy in cattle,poultry and pigs[J]. Animal Health Research Reviews,2008,9(2):201-215.
[11]刘燕坤,罗润波,林焱,等. 噬菌体鸡尾酒对断奶仔猪生长性能、血液参数和粪样菌群的影响[J]. 畜牧兽医学报,2023,54(4):1555-1567.
[12]GEMBARA K, DABROWSKA K. Phage-specific antibodies[J]. Current Opinion in Biotechnology,2021,68:186-192.

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备注/Memo

备注/Memo:
收稿日期:2024-10-24基金项目:“十四五”国家重点研发计划课题项目(2023YFD1800303)作者简介:李星辰(2000-),女,山东潍坊人,硕士研究生,主要从事农产品质量安全研究。(E-mail)lixingchen2000@163.com通讯作者:王冉,(E-mail)ranwang@jaas.ac.cn；何涛,(E-mail)vethetao@163.com
更新日期/Last Update: 2025-07-16