• We are sorry, but NCBI web applications do not support your browser and may not function properly. More information
Logo of jcmPermissionsJournals.ASM.orgJournalJCM ArticleJournal InfoAuthorsReviewers
J Clin Microbiol. Jun 1992; 30(6): 1514–1517.
PMCID: PMC265320

Purification of sputum samples through sucrose improves detection of Mycobacterium tuberculosis by polymerase chain reaction.

Abstract

A method is described for the routine preparation of sputum samples for detection of Mycobacterium tuberculosis by using polymerase chain reaction amplification. Liquefaction of sputum samples with NaOH and subsequent removal of inhibitors of the polymerase reaction with a 50% sucrose centrifugation step (5 min) in a desktop centrifuge allow direct amplification of a 123-bp repetitive region of the M. tuberculosis genome. We have evaluated the sensitivity and specificity of the sucrose method with 155 sputum specimens from patients suspected of having tuberculosis and from normal healthy volunteers. This method, in our opinion, is reproducible, sensitive, and reliable. Multiple samples can be handled simultaneously, and results can be obtained in one working day.

Full text

Full text is available as a scanned copy of the original print version. Get a printable copy (PDF file) of the complete article (834K), or click on a page image below to browse page by page. Links to PubMed are also available for Selected References.

Images in this article

Click on the image to see a larger version.

Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Anargyros P, Astill DS, Lim IS. Comparison of improved BACTEC and Lowenstein-Jensen media for culture of mycobacteria from clinical specimens. J Clin Microbiol. 1990 Jun;28(6):1288–1291. [PMC free article] [PubMed]
  • Böddinghaus B, Rogall T, Flohr T, Blöcker H, Böttger EC. Detection and identification of mycobacteria by amplification of rRNA. J Clin Microbiol. 1990 Aug;28(8):1751–1759. [PMC free article] [PubMed]
  • Brisson-Noël A, Gicquel B, Lecossier D, Lévy-Frébault V, Nassif X, Hance AJ. Rapid diagnosis of tuberculosis by amplification of mycobacterial DNA in clinical samples. Lancet. 1989 Nov 4;2(8671):1069–1071. [PubMed]
  • De Wit D, Steyn L, Shoemaker S, Sogin M. Direct detection of Mycobacterium tuberculosis in clinical specimens by DNA amplification. J Clin Microbiol. 1990 Nov;28(11):2437–2441. [PMC free article] [PubMed]
  • Eisenach KD, Cave MD, Bates JH, Crawford JT. Polymerase chain reaction amplification of a repetitive DNA sequence specific for Mycobacterium tuberculosis. J Infect Dis. 1990 May;161(5):977–981. [PubMed]
  • Gonzalez R, Hanna BA. Evaluation of Gen-Probe DNA hybridization systems for the identification of Mycobacterium tuberculosis and Mycobacterium avium-intracellulare. Diagn Microbiol Infect Dis. 1987 Oct;8(2):69–77. [PubMed]
  • Hance AJ, Grandchamp B, Lévy-Frébault V, Lecossier D, Rauzier J, Bocart D, Gicquel B. Detection and identification of mycobacteria by amplification of mycobacterial DNA. Mol Microbiol. 1989 Jul;3(7):843–849. [PubMed]
  • Hartskeerl RA, de Wit MY, Klatser PR. Polymerase chain reaction for the detection of Mycobacterium leprae. J Gen Microbiol. 1989 Sep;135(9):2357–2364. [PubMed]
  • Hermans PW, Schuitema AR, Van Soolingen D, Verstynen CP, Bik EM, Thole JE, Kolk AH, van Embden JD. Specific detection of Mycobacterium tuberculosis complex strains by polymerase chain reaction. J Clin Microbiol. 1990 Jun;28(6):1204–1213. [PMC free article] [PubMed]
  • Mercier B, Gaucher C, Feugeas O, Mazurier C. Direct PCR from whole blood, without DNA extraction. Nucleic Acids Res. 1990 Oct 11;18(19):5908–5908. [PMC free article] [PubMed]
  • Noordhoek GT, Wieles B, van der Sluis JJ, van Embden JD. Polymerase chain reaction and synthetic DNA probes: a means of distinguishing the causative agents of syphilis and yaws? Infect Immun. 1990 Jun;58(6):2011–2013. [PMC free article] [PubMed]
  • Pao CC, Yen TS, You JB, Maa JS, Fiss EH, Chang CH. Detection and identification of Mycobacterium tuberculosis by DNA amplification. J Clin Microbiol. 1990 Sep;28(9):1877–1880. [PMC free article] [PubMed]
  • Patel RJ, Fries JW, Piessens WF, Wirth DF. Sequence analysis and amplification by polymerase chain reaction of a cloned DNA fragment for identification of Mycobacterium tuberculosis. J Clin Microbiol. 1990 Mar;28(3):513–518. [PMC free article] [PubMed]
  • Pierre C, Lecossier D, Boussougant Y, Bocart D, Joly V, Yeni P, Hance AJ. Use of a reamplification protocol improves sensitivity of detection of Mycobacterium tuberculosis in clinical samples by amplification of DNA. J Clin Microbiol. 1991 Apr;29(4):712–717. [PMC free article] [PubMed]
  • Plikaytis BB, Gelber RH, Shinnick TM. Rapid and sensitive detection of Mycobacterium leprae using a nested-primer gene amplification assay. J Clin Microbiol. 1990 Sep;28(9):1913–1917. [PMC free article] [PubMed]
  • Shankar P, Manjunath N, Mohan KK, Prasad K, Behari M, Shriniwas, Ahuja GK. Rapid diagnosis of tuberculous meningitis by polymerase chain reaction. Lancet. 1991 Jan 5;337(8732):5–7. [PubMed]
  • Sjöbring U, Mecklenburg M, Andersen AB, Miörner H. Polymerase chain reaction for detection of Mycobacterium tuberculosis. J Clin Microbiol. 1990 Oct;28(10):2200–2204. [PMC free article] [PubMed]
  • Thierry D, Brisson-Noël A, Vincent-Lévy-Frébault V, Nguyen S, Guesdon JL, Gicquel B. Characterization of a Mycobacterium tuberculosis insertion sequence, IS6110, and its application in diagnosis. J Clin Microbiol. 1990 Dec;28(12):2668–2673. [PMC free article] [PubMed]
  • Vary PH, Andersen PR, Green E, Hermon-Taylor J, McFadden JJ. Use of highly specific DNA probes and the polymerase chain reaction to detect Mycobacterium paratuberculosis in Johne's disease. J Clin Microbiol. 1990 May;28(5):933–937. [PMC free article] [PubMed]
  • Victor T, du Toit R, van Zyl J, Bester AJ, van Helden PD. Improved method for the routine identification of toxigenic Escherichia coli by DNA amplification of a conserved region of the heat-labile toxin A subunit. J Clin Microbiol. 1991 Jan;29(1):158–161. [PMC free article] [PubMed]

Articles from Journal of Clinical Microbiology are provided here courtesy of American Society for Microbiology (ASM)

Formats:

Related citations in PubMed

See reviews...See all...

Cited by other articles in PMC

See all...

Links

Recent Activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...