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Isolation of a protein antigen from Coxiella burnetii
H P Müller1, N Schmeer, L Rantamäki
1Institute of Hygiene and Infectious Diseases of Animals, Justus-Liebig-Universität, Giessen.
Abstract:
Antigens prepared from Coxiella burnetti, strain Frankfurt, phase II, propagated in persistently infected Buffalo Green Monkey (BGM) cell cultures were purified by guanidinium hydrochloride treatment and chloroform/methanol extraction. By ELISA analysis, chloroform/methanol residues (CMR) proved to be free of host cell antigens. The CMR were sensitive to trypsin, pronase E and proteinase K, as determined by absorption-kinetics of CMR suspensions at 600 nm and release of protein. Coomassie blue stained SDS polyacrylamide gels of proteinase hydrolysates from CMR revealed only a single component of apparently 27,000 D. Silver stained gels, however, showed a second component of apparently 12,000 D. In contrast, from untreated native C. burnetii a large variety of proteins, most of them protease-sensitive, were released by detergents at low temperatures, but the 27,000 D component was only solubilized at 60-100 degrees C. The 27,000 D component was obviously the major protein of CMR as well as of whole cells. Antigenicity of this 27,000 D protein could be demonstrated by agargel precipitation test, ELISA and immunoperoxidase techniques applying antisera raised against whole cells and against the extracted component. The component was also recognized in a dot immunobinding assay by sera from guinea pigs infected with cloned C. burnetii stain Nine Mile, phase I, thus indicating an important role of this antigen in C. burnetii specific immune response.
Insights
Coxiella burnetii antigens were purified, revealing a major 27,000 D protein. This Coxiella burnetii protein is antigenic and plays a key role in the immune response.
Area of Science:
- Microbiology
- Immunology
- Biochemistry
Background:
- Coxiella burnetii is the causative agent of Q fever.
- Antigen preparation is crucial for understanding C. burnetii's immune response.
Purpose of the Study:
- To purify and characterize antigens from Coxiella burnetii.
- To identify major antigenic components and assess their role in immunity.
Main Methods:
- Propagation of C. burnetii in Buffalo Green Monkey (BGM) cells.
- Purification using guanidinium hydrochloride and chloroform/methanol extraction.
- Analysis by ELISA, SDS-PAGE, and various immunological assays (agar-gel precipitation, immunoperoxidase, dot immunobinding).
Main Results:
- Chloroform/methanol residues (CMR) were free of host cell antigens and sensitive to proteases.
- A major 27,000 D protein component was identified in CMR and whole C. burnetii cells.
- This 27,000 D protein demonstrated antigenicity and was recognized by immune sera.
Conclusions:
- The 27,000 D protein is a significant antigen of Coxiella burnetii.
- This protein likely plays a crucial role in the C. burnetii specific immune response.