title: Diagnosis of pulmonary and extrapulmonary tuberculosis based on detection of mycobacterial antigen 85B by immuno-PCR.
abstract: We developed a novel indirect sandwich immuno-polymerase chain reaction (I-PCR) assay for the detection of mycobacterial antigen 85B (Ag85B, 30kDa, Rv1886c) in pulmonary tuberculosis (PTB) and extrapulmonary tuberculosis (EPTB) patients. The amino-modified reporter DNA was covalently attached with the antidetection antibody through a heterobifunctional cross-linking agent succinimidyl 4-[N-maleimidomethyl]-cyclohexane-1-carboxylate. The detection limit of Ag85B by I-PCR was found to be 1 femtogram (fg)/mL, which was 10(6)-fold lower than an analogous enzyme-linked immunosorbent assay (ELISA). The sensitivities of 85% and 77% with I-PCR and 77.6% and 62.5% with ELISA were observed in smear-positive and smear-negative PTB patients, respectively, with high specificity. On the other hand, sensitivities of 84% and 63.7% with I-PCR and 68% and 47.5% with ELISA were observed in confirmed and clinically suspected EPTB cases, respectively, with high specificity.
authors.affiliation: Centre for Biotechnology, Maharshi Dayanand University, Rohtak 124001, Haryana, India.
authors.forename: Netrapal
authors.initials: N
authors.lastname: Singh
authors.affiliation: Department of Biostatistics, All India Institute of Medical Sciences (AIIMS), New Delhi 110029, India.
authors.forename: Vishnubhatla
authors.initials: V
authors.lastname: Sreenivas
authors.affiliation: Department of TB & Respiratory Medicine, University of Health Sciences (UHS), Rohtak 124001, India.
authors.forename: Krishna B
authors.initials: KB
authors.lastname: Gupta
authors.affiliation: Rajan Babu Institute of Pulmonary Medicine and Tuberculosis (RBIPMT), Delhi 110009, India.
authors.forename: Anil
authors.initials: A
authors.lastname: Chaudhary
authors.affiliation: Vallabhbhai Patel Chest Institute (VPCI), University of Delhi, Delhi 110007, India.
authors.forename: Anshu
authors.initials: A
authors.lastname: Mittal
authors.affiliation: Vallabhbhai Patel Chest Institute (VPCI), University of Delhi, Delhi 110007, India.
authors.forename: Mandira
authors.initials: M
authors.lastname: Varma-Basil
authors.affiliation: Vallabhbhai Patel Chest Institute (VPCI), University of Delhi, Delhi 110007, India.
authors.forename: Rajendra
authors.initials: R
authors.lastname: Prasad
authors.affiliation: Centre for Biotechnology, Maharshi Dayanand University, Rohtak 124001, Haryana, India.
authors.forename: Surender K
authors.initials: SK
authors.lastname: Gakhar
authors.affiliation: Department of Biochemistry, Postgraduate Institute of Medical Education and Research, Chandigarh 160012, India.
authors.forename: Gopal K
authors.initials: GK
authors.lastname: Khuller
authors.affiliation: Centre for Biotechnology, Maharshi Dayanand University, Rohtak 124001, Haryana, India. Electronic address: pkmehta3@hotmail.com.
authors.forename: Promod K
authors.initials: PK
authors.lastname: Mehta
mesh_terms: D000217:Acyltransferases / Q000032:analysis*; D000293:Adolescent; D000328:Adult; D000942:Antigens, Bacterial / Q000032:analysis*; D001426:Bacterial Proteins / Q000032:analysis*; D005260:Female; D006801:Humans; D007118:Immunoassay / Q000379:methods*; D008297:Male; D008875:Middle Aged; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D014376:Tuberculosis / Q000175:diagnosis*; D055815:Young Adult
chemical_list: D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D000217:Acyltransferases; C121433:antigen 85B, Mycobacterium tuberculosis
keywords: Antigen 85B; Diagnosis; Immuno-PCR; Mycobacterium tuberculosis; SMCC; Tuberculosis