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. 2016 May 21:16:219.
doi: 10.1186/s12879-016-1552-9.

A novel multi-epitope recombined protein for diagnosis of human brucellosis

Dehui Yin  1 , Li Li  1 , Xiuling Song  1 , Han Li  1   2 , Juan Wang  1 , Wen Ju  1 , Xiaofeng Qu  1 , Dandan Song  1 , Yushen Liu  1 , Xiangjun Meng  1 , Hongqian Cao  1 , Weiyi Song  1 , Rizeng Meng  1   3 , Jinhua Liu  3 , Juan Li  4 , Kun Xu  5
Affiliations

A novel multi-epitope recombined protein for diagnosis of human brucellosis

Dehui Yin et al. BMC Infect Dis. .

Abstract

Background: In epidemic regions of the world, brucellosis is a reemerging zoonosis with minimal mortality but is a serious public hygiene problem. Currently, there are various methods for brucellosis diagnosis, however few of them are available to be used to diagnose, especially for serious cross-reaction with other bacteria.

Method: To overcome this disadvantage, we explored a novel multi-epitope recombinant protein as human brucellosis diagnostic antigen. We established an indirect enzyme-linked immunosorbent assay (ELISA) based on this recombinant protein. 248 sera obtained from three different groups including patients with brucellosis (146 samples), non-brucellosis patients (82 samples), and healthy individuals (20 samples) were tested by indirect ELISA. To evaluate the assay, a receiver-operating characteristic (ROC) analysis and immunoblotting were carried out using these characterized serum samples.

Results: For this test, the area under the ROC curve was 0.9409 (95 % confidence interval, 0.9108 to 0.9709), and a sensitivity of 88.89 % and a specificity of 85.54 % was given with a cutoff value of 0.3865 from this ROC analysis. The Western blot results indicate that it is feasible to differentiate human brucellosis and non-brucellosis with the newly established method based on this recombinant protein.

Conclusion: Our results obtained high diagnostic accuracy of the ELISA assay which encourage the use of this novel recombinant protein as diagnostic antigen to implement serological diagnosis of brucellosis.

Keywords: Brucellosis; Diagnosis; Recombinant protein.

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Figures

Fig. 1
Fig. 1
Amino acid sequence of the recombinant protein in FAST format: “GGGS” is the linker
Fig. 2
Fig. 2
Preparation and identification of recombinant protein. a SDS-PAGE of OMP-induced expression with 0.2 mM IPTG for different times (M, marker; Lane 1, uninduced cells; Lane 2, supernatant of IPTG-induced cells for 2 h; Lane 3, deposition of IPTG-induced cells for 2 h; Lane 4, supernatant of IPTG-induced cells for 4 h; Lane 5, deposition of IPTG-induced cells for 4 h). b SDS-PAGE of OMP purification (M, marker; Lane 1, purified recombinant protein.). c Western blotting analysis of the purified recombinant protein using the anti-His-tag rabbit anti-His-tag polyclonal antibody (M, marker; Lane 1, purified recombinant protein)
Fig. 3
Fig. 3
IELISA analysis of serum samples. The analysis was performed considering positive control serum samples with culture-confirmed/serologically positive brucellosis (146 sera) and negative control sera from patients with other diseases and healthy individuals (102 sera). a Dotplot of the rOMP IELISA assay. b ROC analysis of rOMP IELISA assay results. c Dotplot of the SAT antigen IELISA assay results. d ROC analysis of SAT antigen IELISA assay results
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