Identification, molecular cloning, and evaluation of potential use of isocitrate dehydrogenase II of Mycobacterium bovis BCG in serodiagnosis of tuberculosis.
Diagnosis of tuberculosis is time-consuming and requires infrastructures which are often not available in countries with high incidences of the disease. In the present study, an 82-kDa protein antigen was isolated by affinity chromatography and was identified by peptide mass fingerprinting as isocitrate dehydrogenase II, which is encoded by the icd2 gene of Mycobacterium bovis BCG. The icd2 gene of BCG was cloned by PCR, and the product of recombinant gene expression was purified and analyzed by two-dimensional polyacrylamide gel electrophoresis. The recombinant protein, named rICD2, was tested for its recognition by immunoglobulin G (IgG) antibodies from the sera of 16 patients with tuberculosis (TB) and 23 healthy individuals by Western blotting. The results showed that rICD2 is recognized by IgG antibodies from the sera of all TB patients tested at serum dilutions of > or = 1:640. At a serum dilution of 1:1,280, the sensitivity was 50% and the specificity was 86.9%. These results indicate that rICD2 might represent a candidate for use in a new assay for the serodiagnosis of TB.
Florio
W
W
Dipartimento di Patologia Sperimentale, Biotecnologie Mediche, Infettivologia ed Epidemiologia, Università degli Studi di Pisa, 56127 Pisa, Italy. florio@biomed.unipi.it
Bottai
D
D
Batoni
G
G
Esin
S
S
Pardini
M
M
Maisetta
G
G
Campa
M
M
D000328:Adult; D000368:Aged; D000907:Antibodies, Bacterial / Q000097:blood; D000915:Antibody Affinity; D001426:Bacterial Proteins / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification; D015153:Blotting, Western; D003001:Cloning, Molecular; D005260:Female; D006801:Humans; D007521:Isocitrate Dehydrogenase / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification; D008297:Male; D008875:Middle Aged; D009163:Mycobacterium bovis / Q000201:enzymology / Q000235:genetics / Q000276:immunology*; D012680:Sensitivity and Specificity; D012698:Serologic Tests / Q000379:methods / Q000592:standards; D014376:Tuberculosis / Q000175:diagnosis*
D000907:Antibodies, Bacterial; D001426:Bacterial Proteins; C475629:isocitrate dehydrogenase II, Mycobacterium bovis; D007521:Isocitrate Dehydrogenase