Electroimmunotransferblot assay for the detection of mycobacterialantigens in the cerebrospinal fluid for diagnosis of tubercular meningitis.
BACKGROUND
The diagnosis of tubercular meningitis (TBM) is often delayed as it presents initially with non-specific signs and symptoms leading to delay in appropriate therapy. Conventional modes of diagnosis are time-taking and immunodiagnosis has its own pitfalls. Antigen detection assays have been found to be quite promising in this aspect.


OBJECTIVE
In the present study, attempts were made to evaluate the ElectroImmunoTransferBlot (EITB) test for detection of Mycobacterium tuberculosis antigens in CSF.


METHODS
A total of 46 CSF specimens were collected from 26 clinically suspected cases of TBM and 20 non-TBM cases. The mycobacterial antigens were concentrated by immunoprecipitation and separated based on their molecular weight by SDS-PAGE which were further transferred and immobilized onto a matrix and detected by EITB.


RESULTS
In TBM CSF specimens distinct bands of molecular weight 12 kDa, 30-32 kDa, 71 kDa, 86 kDa, 96 kDa, 110 kDa and 120 kDa were seen in addition to 50 kDa Immunoglobulin (Ig) heavy chain, 25kDa Ig light chain and an indistinct human albumin band at 69kDa. The control group CSF specimens also showed the Ig and albumin bands but showed no cross-reactive antigens. The following proteins 12 kDa (7.7%), 30-32 kDa (23%), 71 kDa (19.2%), 86kDa (77%), 96 kDa (57.5), 110 kDa (23%) and 120 kDa (15.4%) were identified as reactive bands. The results were compared to the reverse passive latex agglutination test.


CONCLUSIONS
The likelihood of diagnosing TBM as evidenced by detecting at least a single mycobacterium specific band was 88.4% by our protocol for antigen detection in CSF. The specificity of EITB for diagnosing TBM was found to be 100% when the 86k Da antigen was excluded from the analysis. However, the method of diagnosis is labour/reagent intensive and needs substantial validation.
Department of Microbiology, Jawaharlal Institute of Postgraduate Medical Education and Research, Pondicherry.
K
K
Venkatesh
Subhash Chandra
SC
Parija
D000942:Antigens, Bacterial / Q000134:cerebrospinal fluid / Q000302:isolation & purification; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D015151:Immunoblotting / Q000379:methods*; D047468:Immunoprecipitation / Q000379:methods*; D009169:Mycobacterium tuberculosis / Q000276:immunology; D012680:Sensitivity and Specificity; D014390:Tuberculosis, Meningeal / Q000175:diagnosis*
D000942:Antigens, Bacterial