Isoelectrophoretic Characterization of Protein Antigens Present in Mycobacterial Culture Filtrates and Recognized by Monoclonal Antibodies Directed Against the <i>Mycobacterium bovis</i> BCG Antigen 85 Complex

Drowart, Annie; Bruyn, Jacqueline De; Huygen, Kris; Damiani, G; Godfrey, Henry P.; Stélandre, Martine; Yernault, Jean Claude; Vooren, J. P. Van

doi:10.1111/j.1365-3083.1992.tb03130.x

Cited by 27 publications

(46 citation statements)

References 16 publications

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“…After completion, proteins were electrophoretically transferred to nitrocellulose sheets. Strips were blocked with fetal calf serum (FCS) and incubated overnight in 1:50 dilution of the pooled serum samples collected at the successive time-points or in 1:10 dilution of monoclonal antibodies (MoAb) directed against the following antigens from M. bovis BCG: 30-32 kDa Ag85 (MoAb TD 32-15 [14,15]); 37-38 kDa PstS-2 (2A-1-2 [16]) and the 65 kDa hsp 65 (1A1 [17]). Antibodies were visualized by peroxidase-conjugated rabbit anti-mouse IgG antibody (Dakopatts, Copenhagen, Denmark) and a-chloronaphthol.…”

Section: Methodsmentioning

confidence: 99%

“…Antibodies in BALB/c sera were predominantly directed against the GroEL-related hsp 65 (and its degradation products) identified by the 1A1 MoAb [17]. Starting from about day 50, additional reactivities against the 37-38 kDa PstS-2 homologue [16], against a 35 kDa protein [13] and against the 30-32 kDa Ag85 complex [14,15] were observed in sera from M. intracellulare-infected BALB/c mice. Sera from M. aviumand M. scrofulaceum-infected BALB/c mice were essentially directed against the hsp 65 stress protein.…”

Section: Antibody Response To Bcg Extract and Culture Filtrate In Micmentioning

confidence: 99%

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Cross‐Reactive Immune Responses Against Mycobacterium bovis BCG in Mice Infected with Non‐Tuberculous Mycobacteria Belonging to the MAIS‐Group

et al. 1997

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Two bacillus Calmette-Guérin (BCG)-susceptible mouse strains, BALB/c and C57BL/6, were infected intravenously with Mycobacterium intracellulare, M. avium or M. scrofulaceum and monitored during 3 months for mycobacterial replication and antibody and Th1-type cytokine production in response to cytoplasmic and secreted antigens from M. bovis BCG. Whereas initial colony-forming unit (CFU) counts of M. intracellulare and M. avium were higher in lungs than in spleen, the opposite was observed for M. scrofulaceum. Mycobacterium intracellulare was the most virulent species and its replication could not be controlled in either mouse strain. It also induced the strongest antibody response. Mycobacterium avium was eliminated in both mouse strains and M. scrofulaceum finally was eliminated in C57BL/6 but multiplied in spleen from BALB/c mice. Significant sustained interleukin-2 and interferon-g production towards BCG antigens was only found in M. scrofulaceum infection. As in BCG-vaccination, M. scrofulaceum-infected C57BL/6 mice demonstrated a higher response towards whole BCG culture filtrate, BCG extract and purified antigen 85 complex (Ag85) from BCG than did BALB/c mice. The data suggest that the presence of M. scrofulaceum in the environment may possibly interfere in genetically predisposed subjects with BCG vaccine and its protective efficacy against M. tuberculosis.

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Section: Methodsmentioning

confidence: 99%

Section: Antibody Response To Bcg Extract and Culture Filtrate In Micmentioning

confidence: 99%

Cross‐Reactive Immune Responses Against Mycobacterium bovis BCG in Mice Infected with Non‐Tuberculous Mycobacteria Belonging to the MAIS‐Group

et al. 1997

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“…Protein expression in the HEK293 cells was verified by Western blotting with anti-His Ab and confirmed further by use of the specific mAb HYB 76-8 to ESAT-6 [41] and TD17 to Ag85B [42]. Large-scale production and purification of virus was done as described previously [43].…”

Section: Viral Constructsmentioning

confidence: 99%

Alteration of epitope recognition pattern in Ag85B and ESAT‐6 has a profound influence on vaccine‐induced protection against Mycobacterium tuberculosis

et al. 2006

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To analyze the effect of vaccine delivery systems on antigen recognition and vaccine efficacy, we compared immune responses in mice immunized either with an adenovirus vector expressing a fusion of Ag85B and ESAT-6 or with the recombinant fusion protein in a liposomal adjuvant. Both vaccines induced high levels of antigen-specific IFN-c production. The adjuvanted protein vaccine induced primarily a CD4 T cell response directed to the epitope Ag85B 241-255 and gave efficient protection against subsequent Mycobacterium tuberculosis infection. In contrast, the adenoviral construct induced a strong CD8 response predominantly targeted to the epitope ESAT-6 15-29 and no significant protection against infection. Vaccination with the protein vaccine resulted in highly accelerated recall of Ag85B 241-255 -specific T cells immediately post M. tuberculosis challenge whereas the ESAT-6 15-29 epitope was barely recognized during infection. Delivery of the viral construct in cationic liposomes switched the immune response to a protective one dominated by CD4 T cells targeted to the Ag85B [241][242][243][244][245][246][247][248][249][250][251][252][253][254][255] epitope. These data demonstrate that the nature of the T cell response to a vaccine antigen is more important than its magnitude with respect to protective efficacy and that vaccine-mediated changes in immunodominance can result in T cell responses of limited relevance during the natural infection.

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“…Unitage for each subsequent antigen preparation was determined by parallel-line analysis of dot blots of initial and subsequent materials (7). Mouse immunoglobulin G1 (IgG 1 ) anti-BCG Ag85 complex, clone 240 (7) (14). The Ag85 epitope recognized by clone 240 is not known but is probably conformational (reference 21 and our unpublished observations).…”

Section: Methodsmentioning

confidence: 99%

Combinatorial Use of Antibodies to Secreted Mycobacterial Proteins in a Host Immune System-Independent Test for Tuberculosis

et al. 2001

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Laboratory diagnosis of tuberculosis is often difficult. Immunodetection of circulating Mycobacterium tuberculosis proteins shed during active infection would not depend on an intact host immune response and could take advantage of the speed and low costs afforded by antibody-based assays. We previously showed that patients with active tuberculosis had increased levels of circulating antigen 85 (Ag85) proteins independent of their tuberculin skin test status (S. I. Bentley-Hibbert, X. Quan, T. Newman, K. Huygen, and H. P. Godfrey, Infect. Immun. 67:581-588, 1999). To extend these observations to a Mycobacterium bovis BCG-vaccinated population and to another secreted mycobacterial protein, Ag85 and PstS-1 (protein antigen B, p38 antigen) were quantified in sera from 97 Chilean tuberculosis patients and healthy controls (many of whom had received BCG as children) using dot immunobinding, mouse monoclonal anti-BCG Ag85 complex antibody, and chicken antipeptide antibodies reactive with M. tuberculosis Ag85B and PstS-1. The latter antibodies had been raised to peptide-derived immunogens expressed on a novel proprietary protein carrier in Escherichia coli. Median serum Ag85 levels measured by using either anti-Ag85 antibody were significantly higher in patients with active tuberculosis than in healthy controls (P, <0.001 to 0.01); the median serum PstS-1 levels were similar in patients and controls. The sensitivity of significantly elevated circulating Ag85 levels in patients with pulmonary tuberculosis measured by anti-Ag85 complex or anti-Ag85B antibodies was 60 and 55%, respectively, but increased to 77% when results obtained with both anti-Ag85 antibodies were considered jointly (P < 0.02). The corresponding specificities for individual and joint consideration were 95, 85, and 80%, respectively. These results indicate that elevated Ag85 levels can be detected in patients with active tuberculosis even after BCG vaccination and suggest that combinatorial use of antibodies directed at different epitopes of this protein could provide a viable strategy for developing new host immune response-independent diagnostic tests for tuberculosis.

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Isoelectrophoretic Characterization of Protein Antigens Present in Mycobacterial Culture Filtrates and Recognized by Monoclonal Antibodies Directed Against the Mycobacterium bovis BCG Antigen 85 Complex

Cited by 27 publications

References 16 publications

Cross‐Reactive Immune Responses Against Mycobacterium bovis BCG in Mice Infected with Non‐Tuberculous Mycobacteria Belonging to the MAIS‐Group

Cross‐Reactive Immune Responses Against Mycobacterium bovis BCG in Mice Infected with Non‐Tuberculous Mycobacteria Belonging to the MAIS‐Group

Alteration of epitope recognition pattern in Ag85B and ESAT‐6 has a profound influence on vaccine‐induced protection against Mycobacterium tuberculosis

Combinatorial Use of Antibodies to Secreted Mycobacterial Proteins in a Host Immune System-Independent Test for Tuberculosis

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