EVALUATION OF A PCR ASSAY FOR DETECTION OF MYCOBACTERIUM-TUBERCULOSISIN CLINICAL SPECIMENS

A polymerase chain reaction (PCR) assay for detection of M. tuberculos is was optimized for application to clinical specimens, which were pre pared for amplification by boiling in buffer. The buffer contained a s ynthetic DNA fragment to determine if DNA amplification from the indiv idual prepared specimens was subject to inhibition because of substanc es present in the specimen, or by the process of specimen preparation or storage. The PCR test was less sensitive than direct microscopy (75 % as against 87.5%) and had a specificity of 97%. Invalid results due to inhibition of amplification occurred in 12% of specimens. Incorpora tion of the internal standard into the specimen preparation buffer ens ures that any step in the process which inhibits DNA amplification is detected in the failure of amplification of the internal standard. The use of internal standard in this way should be considered in developi ng diagnostic protocols.