SCANNING ELECTRON-MICROSCOPIC METHOD FOR INTERNAL STRUCTURE-ANALYSIS OF ANAEROBIC GRANULAR SLUDGE

The development and disintegration of anaerobic granular sludge are crucial factors for successful application of anaerobic wastewater treatment using Upflow Anaerobic Sludge Bed (UASB) reactors. Knowledge of the internal structure of anaerobic sludge granules is essential for understanding these processes. The scanning electron microscope is widely used in biofilm research because of the 3 dimensional view. However, sectioning is necessary for internal structure research but induces severe artifacts. In this study a method was developed to minimize sectioning artifacts during preparation for scanning electron microcopy research on anaerobic granular sludge. Cryo-fixation was used to increase the mechanical strength of the samples. To avoid ice crystal damage and to improves the sectioning result, DMSO and ethanol were used as cryo-protectants. Best results were obtained using DMSO as a cryoprotectant. Smooth section surface areas were obtained using 50% DMSO-treated, fast frozen granules sectioned at 110 degrees C.