Full-Text Search:
Home|Journal Papers|About CNKI|User Service|FAQ|Contact Us|中文
《Progress in Veterinary Medicine》 2017-12
Add to Favorite Get Latest Update

Prokaryotic Expression of HA Protein of Haemophilus paragallinarum Serotype A and Establishment and Preliminary Application of Indirect ELISA Detection Method

ZHANG Man;HAN Fei;College of Animal Engineering,Yangling Vocational Technical College;  
After prokaryotic expression of Haemophilus paragallinarum serotype A HA protein,and using the HA protein as the antigen,the indirect ELISA detection method was established.Hpgserotype A HA protein gene was cloned into E.coli expression vector pET-28 a(+)in this study to induce the expression of recombinant HA protein.HA protein was purified and tested by Western blot.Using the recombinant HA protein as the antigen,the indirect ELISA detection method was established to detect the level of Hpg antibody in serum.Specificity,sensitivity and reproducibility tests were carried out,and the clinical samples were tested.The recombinant protein with the size of 37 ku was obtained.Western blot showed that the purified HA protein had good specificity and antigen reactivity.The optimal reaction conditions were established as follows:the coating amount of antigen was 500 ng per hole,closed 1 h;the serum dilution was1∶200,reacting 1 h;the second antibody dilution was 1∶2 500,reacting 1 h;the TMB reacting time was10 min.Criteria for determining the negative and positive results of serum samples were OD_(450)≥0.34 and OD_(450)0.34,respectively.Specificity tests showed that the results of Newcastle disease virus,avian influenza virus,Salmonella spp,Staphylococcus aureus,Pasteurella multocida,Escherichia coli positive sera were negative.85 positive sera were tested and the sensitivity was 98.8% .The repeatability test showed that the coefficient of variation of OD_(450) between the same batch and different batch were 1.5% -3.7% and 6.5% -8% ,respectively.The clinical 215 serum samples were tested using established system and the positive rate was 25.1% .The established system in this test can be used for clinical detection of serum antibody and epidemiological investigation of Hpgserotype A.
【Fund】: 陕西省科技攻关项目(2016NY-096);; 杨凌示范区重大科技项目(2016-XY-16)
【CateGory Index】: S852.61
Download(CAJ format) Download(PDF format)
CAJViewer7.0 supports all the CNKI file formats; AdobeReader only supports the PDF format.
©2006 Tsinghua Tongfang Knowledge Network Technology Co., Ltd.(Beijing)(TTKN) All rights reserved