Abstract
Analysis by Western immunoblotting of Mycobacterium bovis BCG short-term culture filtrates with a pool of serum samples from lepromatous leprosy patients revealed an immunodominant protein doublet with apparent molecular masses of 28 and 30 kilodaltons (kDa). The humoral response to these antigens was also investigated by using individual serum samples from patients representative of the whole spectrum of leprosy and from tuberculosis patients, as well as from contacts of leprosy patients and control groups. The protein doublet was recognized by 92% of the sera from patients with lepromatous leprosy (51 of 56), whereas essentially negative results were obtained with sera from the other groups. Similar immunodominant bands were also detected by Western blotting analysis of sonic extracts of seven other slow- and fast-growing mycobacterial species, suggesting a broad distribution of these antigens within the genus. Analysis of the purified doublet by Western blotting after two-dimensional gel electrophoresis fractionation showed that the 28- and 30-kDa doublet consisted of at least five different components with pls from 5.2 to 5.7 and molecular masses from 28 to 31 kDa. These results indicate that the protein doublet could be used as a potential marker in the diagnosis of lepromatous leprosy.
Full text
PDF![2184](https://cdn.ncbi.nlm.nih.gov/pmc/blobs/d05c/266990/806e97108d0f/jcm00070-0052.png)
![2185](https://cdn.ncbi.nlm.nih.gov/pmc/blobs/d05c/266990/d7d07c839cfa/jcm00070-0053.png)
![2186](https://cdn.ncbi.nlm.nih.gov/pmc/blobs/d05c/266990/d96cd98a4101/jcm00070-0054.png)
![2187](https://cdn.ncbi.nlm.nih.gov/pmc/blobs/d05c/266990/6dc451b60b2e/jcm00070-0055.png)
![2188](https://cdn.ncbi.nlm.nih.gov/pmc/blobs/d05c/266990/4441e6c51aa8/jcm00070-0056.png)
![2189](https://cdn.ncbi.nlm.nih.gov/pmc/blobs/d05c/266990/84e3de39400e/jcm00070-0057.png)
Images in this article
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Abou-Zeid C., Smith I., Grange J. M., Ratliff T. L., Steele J., Rook G. A. The secreted antigens of Mycobacterium tuberculosis and their relationship to those recognized by the available antibodies. J Gen Microbiol. 1988 Feb;134(2):531–538. doi: 10.1099/00221287-134-2-531. [DOI] [PubMed] [Google Scholar]
- Britton W. J., Hellqvist L., Garsia R. J., Basten A. Antigens of Mycobacterium leprae identified by immunoprecipitation with sera from leprosy and tuberculosis patients. Clin Exp Immunol. 1988 Mar;71(3):394–398. [PMC free article] [PubMed] [Google Scholar]
- Closs O., Harboe M., Axelsen N. H., Bunch-Christensen K., Magnusson M. The antigens of Mycobacterium bovis, strain BCG, studied by crossed immunoelectrophoresis: a reference system. Scand J Immunol. 1980;12(3):249–263. doi: 10.1111/j.1365-3083.1980.tb00065.x. [DOI] [PubMed] [Google Scholar]
- Collins F. M., Lamb J. R., Young D. B. Biological activity of protein antigens isolated from Mycobacterium tuberculosis culture filtrate. Infect Immun. 1988 May;56(5):1260–1266. doi: 10.1128/iai.56.5.1260-1266.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Daniel T. M., Ferguson L. E. Purification and Characterization of Two Proteins from Culture Filtrates of Mycobacterium tuberculosis H(37)Ra Strain. Infect Immun. 1970 Feb;1(2):164–168. doi: 10.1128/iai.1.2.164-168.1970. [DOI] [PMC free article] [PubMed] [Google Scholar]
- De Bruyn J., Bosmans R., Turneer M., Weckx M., Nyabenda J., Van Vooren J. P., Falmagne P., Wiker H. G., Harboe M. Purification, partial characterization, and identification of a skin-reactive protein antigen of Mycobacterium bovis BCG. Infect Immun. 1987 Jan;55(1):245–252. doi: 10.1128/iai.55.1.245-252.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Harboe M., Closs O., Bjorvatn B., Kronvall G., Axelsen N. H. Antibody response in rabbits to immunization with Mycobacterium leprae. Infect Immun. 1977 Dec;18(3):792–805. doi: 10.1128/iai.18.3.792-805.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Hunter S. W., Gaylord H., Brennan P. J. Structure and antigenicity of the phosphorylated lipopolysaccharide antigens from the leprosy and tubercle bacilli. J Biol Chem. 1986 Sep 15;261(26):12345–12351. [PubMed] [Google Scholar]
- Huygen K., Van Vooren J. P., Turneer M., Bosmans R., Dierckx P., De Bruyn J. Specific lymphoproliferation, gamma interferon production, and serum immunoglobulin G directed against a purified 32 kDa mycobacterial protein antigen (P32) in patients with active tuberculosis. Scand J Immunol. 1988 Feb;27(2):187–194. doi: 10.1111/j.1365-3083.1988.tb02338.x. [DOI] [PubMed] [Google Scholar]
- Kaufmann S. H., Chiplunkar S., Flesch I., De Libero G. Possible role of helper and cytolytic T cells in mycobacterial infections. Lepr Rev. 1986 Dec;57 (Suppl 2):101–111. doi: 10.5935/0305-7518.19860060. [DOI] [PubMed] [Google Scholar]
- Klatser P. R., van Rens M. M., Eggelte T. A. Immunochemical characterization of Mycobacterium leprae antigens by the SDS-polyacrylamide gel electrophoresis immunoperoxidase technique (SGIP) using patients' sera. Clin Exp Immunol. 1984 Jun;56(3):537–544. [PMC free article] [PubMed] [Google Scholar]
- LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
- Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
- Løvik M., Closs O. Induction of immunity against live Mycobacterium lepraemurium: a requirement for viable bacilli? Immunology. 1984 Sep;53(1):165–173. [PMC free article] [PubMed] [Google Scholar]
- Matsuo K., Yamaguchi R., Yamazaki A., Tasaka H., Yamada T. Cloning and expression of the Mycobacterium bovis BCG gene for extracellular alpha antigen. J Bacteriol. 1988 Sep;170(9):3847–3854. doi: 10.1128/jb.170.9.3847-3854.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Müller I., Cobbold S. P., Waldmann H., Kaufmann S. H. Impaired resistance to Mycobacterium tuberculosis infection after selective in vivo depletion of L3T4+ and Lyt-2+ T cells. Infect Immun. 1987 Sep;55(9):2037–2041. doi: 10.1128/iai.55.9.2037-2041.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Nagai S., Matsumoto J., Nagasuga T. Specific skin-reactive protein from culture filtrate of Mycobacterium bovis BCG. Infect Immun. 1981 Mar;31(3):1152–1160. doi: 10.1128/iai.31.3.1152-1160.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
- O'Farrell P. H. High resolution two-dimensional electrophoresis of proteins. J Biol Chem. 1975 May 25;250(10):4007–4021. [PMC free article] [PubMed] [Google Scholar]
- Ratliff T. L., McGarr J. A., Abou-Zeid C., Rook G. A., Stanford J. L., Aslanzadeh J., Brown E. J. Attachment of mycobacteria to fibronectin-coated surfaces. J Gen Microbiol. 1988 May;134(5):1307–1313. doi: 10.1099/00221287-134-5-1307. [DOI] [PubMed] [Google Scholar]
- Ridley D. S., Jopling W. H. Classification of leprosy according to immunity. A five-group system. Int J Lepr Other Mycobact Dis. 1966 Jul-Sep;34(3):255–273. [PubMed] [Google Scholar]
- Rook G. A. The immunogenicity of killed mycobacteria. Lepr Rev. 1980 Dec;51(4):295–301. doi: 10.5935/0305-7518.19800030. [DOI] [PubMed] [Google Scholar]
- Rumschlag H. S., Shinnick T. M., Cohen M. L. Serological responses of patients with lepromatous and tuberculoid leprosy to 30-, 31-, and 32-kilodalton antigens of Mycobacterium tuberculosis. J Clin Microbiol. 1988 Oct;26(10):2200–2202. doi: 10.1128/jcm.26.10.2200-2202.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Steinhoff U., Kaufmann S. H. Specific lysis by CD8+ T cells of Schwann cells expressing Mycobacterium leprae antigens. Eur J Immunol. 1988 Jun;18(6):969–972. doi: 10.1002/eji.1830180622. [DOI] [PubMed] [Google Scholar]
- Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Wiker H. G., Harboe M., Lea T. E. Purification and characterization of two protein antigens from the heterogeneous BCG85 complex in Mycobacterium bovis BCG. Int Arch Allergy Appl Immunol. 1986;81(4):298–306. doi: 10.1159/000234153. [DOI] [PubMed] [Google Scholar]
- Wiker H. G., Harboe M., Nagai S., Patarroyo M. E., Ramirez C., Cruz N. MPB59, a widely cross-reacting protein of Mycobacterium bovis BCG. Int Arch Allergy Appl Immunol. 1986;81(4):307–314. doi: 10.1159/000234154. [DOI] [PubMed] [Google Scholar]
- Young D. B. Structure of mycobacterial antigens. Br Med Bull. 1988 Jul;44(3):562–583. doi: 10.1093/oxfordjournals.bmb.a072268. [DOI] [PubMed] [Google Scholar]
- Zinkernagel R. M., Doherty P. C. MHC-restricted cytotoxic T cells: studies on the biological role of polymorphic major transplantation antigens determining T-cell restriction-specificity, function, and responsiveness. Adv Immunol. 1979;27:51–177. doi: 10.1016/s0065-2776(08)60262-x. [DOI] [PubMed] [Google Scholar]