Utility of dominant epitopes derived from cell-wall protein LppZ for immunodiagnostic of pulmonary tuberculosis

Abstract

Background: Serological antibodies tests for tuberculosis (TB) are widely used in developing countries. They appear to have some advantages- faster, simple and could be used for extrapulmonary TB. However, most of current commercial TB serological tests are failed to provide sufficient sensitivity and specificity. Improved serological biomarkers were essential. In this study, we present an approach using peptide array to discover new immunodiagnostic biomarkers based on immunodominant epitopes of TB antigens.

Results: The Probable conserved lipoprotein LppZ, which is difficult to express and purify in vivo was selected as the model antigen. We use two-step screening for dominant epitope selection. Based on peptide array data from 170 TB patients and 41 control samples, two dominant epitopes were identified to have diagnostic value for TB patients. Truncation assay was used to identify the core reactive sequence. Peptide- based ELISA was used to evaluate the diagnostic ability of pep-LppZ-1 and pep-LppZ-13. Pep-LppZ-1 has a sensitivity of 49.2% and a specificity of 83.3% in TB diagnose. Pep-LppZ-13 has a sensitivity of 43.3% and a specificity of 88.5% in TB diagnose.

Conclusions: Our result demonstrated that peptide array screening would be an advantage strategy of screening TB diagnostic peptides.

Keywords: LppZ; Serologic test; Tuberculosis peptide arrays.

Fig. 1

Screening for LppZ dominated epitopes. a Second round peptide array screen of 16 candidate epitope peptides selected from first round screening. J091 to J140 stands for serum sample name; CT represents positive control. b Scattergram of peptide arrays signals. Data were normalized by CT signal on each array. Full-length blots are presented in Additional file 1: Figure S1

Fig. 2

Identify the core sequence of candidate peptides during serum antibody reaction. A: Truncation assay on pep-LppZ-1 and pep-Lppz-13 peptides. N➡ indicates the truncation starts at the N terminal; C ➡ indicates the truncation starts at the C terminal. WT stands for wide type. Each letter means the amino acid was removed. Full-length blots are presented in Additional file 1: Figure S2

Fig. 3

Validate and evaluate the reactivity of dominated epitopes by ELISA. a Histogram of relative concentration of antibodies toward dominated epitopes. b & c: ROC (receiver operating characteristic) curve of pep-LppZ-1 and pep-LppZ-13 on detecting TB infection based on ELISA data