Assessment of the IgA immunoassay diagnostic potential of the Mycobacterium tuberculosis MT10.3-MPT64 fusion protein in tuberculous pleural fluid.
Pleural tuberculosis (PL-TB) remains difficult to diagnose. An enzyme-linked immunosorbent assay (ELISA) was developed based on a construction containing the fusion of the Rv3019c (MT10.3) and Rv1980c (MPT64) gene sequences, and its performance was evaluated in an area where TB is endemic. A total of 92 pleural fluid (PF) samples at serial dilutions of 1:50 to 1:800 were included in the ELISA IgA MT10.3-MPT64 evaluation: 70 from TB patients and 22 from patients with other pleurisies. Confirmation of the expression and subsequent purification of the protein was made by SDS-PAGE and Western blot assays, resulting in a 36-kDa protein. ELISA IgA MT10.3-MPT64 showed sensitivities of 61.4%, 58.6%, 62.9%, 67.1%, and 70% at each PF dilution, respectively. The cumulative results of all dilutions increased sensitivity to 81.4% without jeopardizing specificity. Similar results were also obtained at the combined dilutions of 1:50, 1:200, and 1:800 or 1:50 plus 1:800 dilutions (80%). The overall sensitivity of the reference test, i.e., histopathological examination, was 74%. But, via the ELISA IgA MT10.3-MPT64 test, sensitivity was high for specimens with a negative culture (23/27; 85.2%) or nonspecific histopathology (17/18; 94.4%). Our findings demonstrated the promising use of this test as an adjunct in PL-TB diagnoses, particularly in cases with lower bacterial loads and false-negative results in the reference tests, since the new test includes such important features as quick and easy application, high sensitivity and, perhaps most importantly, affordability, which is so crucial for its widespread use in developing countries.
Laboratory of Cellular Microbiology, Oswaldo Cruz Institute, Fiocruz, Rio de Janeiro, Brazil.
Leonardo Silva
LS
Araujo
Renata de Moraes
RM
Maciel
Renata Maciel
RM
Moraes
Anete
A
Trajman
Maria Helena Féres
MH
Saad
D000293:Adolescent; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000907:Antibodies, Bacterial / Q000032:analysis*; D000942:Antigens, Bacterial / Q000235:genetics / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D005260:Female; D006801:Humans; D007070:Immunoglobulin A / Q000032:analysis*; D008297:Male; D008875:Middle Aged; D009169:Mycobacterium tuberculosis / Q000276:immunology*; D010996:Pleural Effusion / Q000276:immunology*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology; D012680:Sensitivity and Specificity; D014396:Tuberculosis, Pleural / Q000175:diagnosis*; D055815:Young Adult
D000907:Antibodies, Bacterial; D000942:Antigens, Bacterial; D007070:Immunoglobulin A; C510537:MPT64 protein, Mycobacterium tuberculosis; D011993:Recombinant Fusion Proteins