title: Development of an ultrasensitive polymerase chain reaction-amplified immunoassay based on mycobacterial RD antigens: implications for the serodiagnosis of tuberculosis.
abstract: Immuno-polymerase chain reaction (I-PCR) combines the versatility of enzyme-linked immunosorbent assay (ELISA) with the exponential amplification power of PCR. The present study was designed to detect antibodies to Mycobacterium tuberculosis complex-specific region of difference (RD) antigens, i.e., early secretory antigenic target-6, culture filtrate protein-10, culture filtrate protein-21, and mycobacterial protein from species tuberculosis-64, as well as antigens in pulmonary tuberculosis patients by I-PCR assay. We could detect ESAT-6 and other RD antigens up to 0.1 fg by I-PCR assay, thus resulting in 10(7) times higher sensitivity than that observed with ELISA. With paired sample analysis based on the detection of antibodies in serum and antigens in sputum of the same individual, the sensitivity of RD multi-antigen cocktail-based I-PCR assay was 72% in smear-negative cases and 91% in smear-positive cases of pulmonary tuberculosis with high specificity values. In extrapulmonary tuberculosis patients, higher sensitivity was observed by detecting cocktail of antigens by I-PCR assay as compared to sensitivity earlier observed in the same samples by ELISA.
authors.affiliation: Centre for Biotechnology, Maharshi Dayanand University, Rohtak-124001, India.
authors.forename: Promod K
authors.initials: PK
authors.lastname: Mehta
authors.forename: Mamta
authors.initials: M
authors.lastname: Kalra
authors.forename: Gopal Krishan
authors.initials: GK
authors.lastname: Khuller
authors.forename: Digambar
authors.initials: D
authors.lastname: Behera
authors.forename: Indu
authors.initials: I
authors.lastname: Verma
mesh_terms: D000907:Antibodies, Bacterial / Q000097:blood*; D000942:Antigens, Bacterial / Q000032:analysis* / Q000235:genetics / Q000276:immunology; D016022:Case-Control Studies; D004269:DNA, Bacterial / Q000032:analysis; D006801:Humans; D007118:Immunoassay / Q000379:methods*; D009169:Mycobacterium tuberculosis / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D013183:Sputum / Q000276:immunology / Q000382:microbiology; D014376:Tuberculosis / Q000097:blood / Q000175:diagnosis* / Q000276:immunology
chemical_list: D000907:Antibodies, Bacterial; D000942:Antigens, Bacterial; D004269:DNA, Bacterial