Immunoproteomic Analysis Of Mycobacterium Tuberculosis And Nontuberculous Mycobacteria Towards The Discovery Of Potential Universal Or Species Specific Biomarkers
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Date
2017-01
Authors
Razman, Aimi Zafira
Journal Title
Journal ISSN
Volume Title
Publisher
Universiti Sains Malaysia
Abstract
The immunoproteomic analysis of whole cell protein (WCP) of Mycobacterium
tuberculosis complex (MTBC) and non–tuberculous mycobacteria (NTM) was carried
out to identify the potential species specific and cross genus conserved species
biomarkers present in the mycobacterial species. The WCP of Mycobacterium species
were separated using SDS-PAGE and followed by Western blot assay to detect IgM,
IgG and IgA immune response in pooled sera of TB patients and healthy subjects as
well as the individual sera. The immunoreactive bands in WB assay were divided into
4 region of antigens; (1) above 67 kDa; (2) 67 kDa – 32 kDa; (3) 30 kDa, 28kDa and
26 kDa; and (4) below 25 kDa. Based on the WB results, the two species selected for
further investigation were M. tuberculosis H37Rv and M. abscessus. The WCP of
these species were purified and fractionated by preparative rotofor isoelectric focusing
(IEF) system and characterized by analytical polyacrylamide gel (PAGE IEF) followed
by immunoblotting. Based on the immunoblotting results, 2 fractions from each
species were selected. These include fraction F1 and F16 of M. tuberculosis H37Rv
and fraction F1 and F3 of M. abscessus. The proteins from the selected fractions were
subjected to identification by MALDI-TOF MS. Employing the approach, a total of 4
bands were excised from preparative PAGE-IEF gel of M. tuberculosis H37Rv’s
fractions (encoded as: T1B1, T1B2, T16B1 and T16B2) and a total of 7 bands from
M. abscessus’s fractions (encoded as: A1B1, A1B2, A2B3, A3B1, A3B2, A3B3 and
A3B4). The MALDI-TOF led to the identification of 21 peptides from M. tuberculosis
H37Rv and 8 peptides from M. abscessus.
Description
Keywords
Immunoproteomic analysis of Mycobacterium tuberculosis , the discovery of species specific biomarkers