Immuno-electron microscopy has allowed the selective localisation of molecules with high resolution and high specificity. Cryopreparatory methods have provided better retention of antigenicity suitable for precise immunolabelling together with optimal structural preservation of cellular components. Cryosubstitution and cryoultramicrotomy have widely been exploited for immunolabelling. Molecular Distillation Dryer (MDD), a form of freeze-drying technique, has recently been used for immunolabelling of Plasmodium falciparum stress proteins and nuclear ribonucleoprotein particles in cultured cells. In the present study, we report the comparison of all three cryotechniques in the immunolabelling of bacterial antigens of Coxiella burnetii.The highly infectious C. burnetii was prefixed in 3% glutaraldehyde (66 mM cacodylate buffer, pH 6.8 ). The cells were then pre-embedded in 2% low-temperature agarose on Durapore hydrophilic membrane prior to cryofixation using a LifeCell CF100 metal-mirror system. A 1% glutaraldehyde in 100% methanol was used as a medium for cryosubstitution in a Reichert CS Auto Cryosubstitution apparatus.
Multiple Substrate Usage of Coxiella burnetii to Feed a Bipartite Metabolic Network
