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Characterization of PPD protein antigens in whole cell lysates of Mycobacterium bovis BCG
S S Bardarov1, J Kriakov, A Karakashyan
1Department of Molecular Biology, Bulgarian Academy of Sciences, Sofia.
Abstract:
The low molecular mass protein antigens in PPD from M. bovis BCG were chemically oligomerized using sulfosuccinimidyl-4-(p-maleimidophenyl)-butyrate (S-SMPB) as a crosslinking agent. Protein oligomers with molecular mass over 90 kDa were obtained and used for the preparation of hyperimmune polyclonal rabbit antiserum. Using this antiserum four protein bands with molecular mass 120, 90, 75 and 65 kDA were detected in immunoblotting analysis of sonic extract from M. bovis BCG separated in SDS-polyacrylamide gel. We suggest that these immunoreactive proteins in the sonic extract represent the native forms of the heat stable low molecular mass protein antigens in PPD.
Insights
Researchers chemically oligomerized low molecular mass protein antigens from Mycobacterium bovis bacille Calmette-Guérin purified protein derivative (PPD). This process identified four native heat-stable protein antigens in M. bovis BCG PPD.
Area of Science:
- Immunology
- Microbiology
- Biochemistry
Background:
- Purified protein derivative (PPD) from Mycobacterium bovis bacille Calmette-Guérin (BCG) contains low molecular mass protein antigens.
- Characterizing these antigens is crucial for understanding immune responses to M. bovis BCG.
Purpose of the Study:
- To chemically oligomerize low molecular mass protein antigens from M. bovis BCG PPD.
- To generate hyperimmune polyclonal rabbit antiserum against these oligomers.
- To identify potential native forms of these antigens in M. bovis BCG sonic extracts.
Main Methods:
- Chemical oligomerization of low molecular mass protein antigens using sulfosuccinimidyl-4-(p-maleimidophenyl)-butyrate (S-SMPB).
- Preparation of hyperimmune polyclonal rabbit antiserum against the obtained protein oligomers.
- Immunoblotting analysis of M. bovis BCG sonic extract using the developed antiserum.
Main Results:
- Protein oligomers with molecular mass exceeding 90 kDa were successfully obtained.
- Immunoblotting detected four distinct protein bands (120, 90, 75, and 65 kDa) in M. bovis BCG sonic extract.
- These immunoreactive proteins are suggested to be the native forms of the heat-stable low molecular mass antigens in PPD.
Conclusions:
- The study successfully identified and characterized potential native protein antigens within M. bovis BCG PPD.
- The findings contribute to a better understanding of the antigenic composition of M. bovis BCG.
- These identified proteins may serve as targets for improved diagnostics or vaccines.