This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Fiallo, P Articles by Gillis, T P Search for Related Content PubMed PubMed Citation Articles by Fiallo, P Articles by Gillis, T P

Effects of fixation on polymerase chain reaction detection of Mycobacterium leprae.

CIR LEP, University of Genoa, Italy.

ABSTRACT

The effects of standard fixatives (10% neutral buffered formalin, ethanol and mercury based) on the detection of Mycobacterium leprae DNA by the polymerase chain reaction (PCR) were studied. Mercury-based fixatives (Zenker's and Carnoy-Lebrun's fluids) strongly inhibited PCR amplification of M. leprae DNA. Ten percent neutral buffered formalin was inhibitory, but significant inhibition was observed only when fixation times exceeded 24 h. Ethanol-based fixatives provided the best medium for holding specimens for subsequent PCR with both free bacilli and skin biopsy specimens containing M. leprae. The M. leprae-specific, 360-bp region of the 18-kDa protein gene could be amplified from paraffin-embedded sections of formalin-fixed skin biopsy specimens from patients with either multibacillary or paucibacillary infections when proper fixation conditions were used. Results of the study demonstrate that tissues properly fixed with two standard fixatives (10% neutral buffered formalin and 50 or 70% ethanol) can be analyzed by PCR for the presence of M. leprae with no loss in specificity and only minimal diminution in sensitivity compared with the specificities and sensitivities obtained by use of freshly prepared, unfixed specimens.

This article has been cited by other articles:

• Ortega, N., Navarro, J. A., Nicolas, L., Buendia, A. J., Caro, M. R., Del Rio, L., Martinez, C. M., Cuello, F., Salinas, J., Gallego, M. C. (2007). Evaluation of Chlamydophila abortus DNA extraction protocols for polymerase chain reaction diagnosis in paraffin-embedded tissues. jvdi 19: 421-425 [Abstract] [Full Text]  
• Scollard, D. M., Adams, L. B., Gillis, T. P., Krahenbuhl, J. L., Truman, R. W., Williams, D. L. (2006). The Continuing Challenges of Leprosy. Clin. Microbiol. Rev. 19: 338-381 [Abstract] [Full Text]  
• Williams, D. L., Pittman, T. L., Gillis, T. P., Matsuoka, M., Kashiwabara, Y. (2001). Simultaneous Detection of Mycobacterium leprae and Its Susceptibility to Dapsone Using DNA Heteroduplex Analysis. J. Clin. Microbiol. 39: 2083-2088 [Abstract] [Full Text]  
• DONOGHUE, H.D., HOLTON, J., SPIGELMAN, M. (2001). PCR primers that can detect low levels of Mycobacterium leprae DNA. J Med Microbiol 50: 177-182 [Abstract] [Full Text]  
• SALIAN, N. V., RISH, J. A., EISENACH, K. D., DONALD CAVE, M., BATES, J. H. (1998). Polymerase Chain Reaction to Detect Mycobacterium tuberculosis in Histologic Specimens. Am. J. Respir. Crit. Care Med. 158: 1150-1155 [Abstract] [Full Text]