A Simple Method for Extraction of Lipopolysaccharides from Brucella Melitensis

Brucellosis is the most common zoonotic bacterial disease around the world, and its causative agent is Brucella, a gram-negative bacterium. Lipopolysaccharide (LPS) molecules are the most significant surface antigen of Brucella bacteria. This antigen is used in a high number of research and diagnostic fields. The present study was designed in order to introduce a simple method to extract LPS from Brucella melitensis . Materials and Meth- ods: S mooth strains of B. melitensis were cultured in large quantity, and LPS was extracted through changes in the hot-phenol method. The LPS concentration was measured by using the dimethyl-methylene blue method. The contamination rates of proteins and nucleic acids were determined using the bicinchoninic acid (BCA) method and absorbance measurement at 260 nm, respectively.

The amount of the extracted LPS was 1.1% of the wet weight of the bacteria. The rate of contamination with nucleic acid was measured to be 0.2% of the LPS. The protein contamination was not detectable through the BCA method.

The advantages of our newly-introduced method are considerable decrease in the time required to extract LPS; independence from nucleic acid and protein digestive enzymes; an extraction rate of LPS equivalent to other applied methods; nucleic acid contamination equal or lower than other methods, and no protein contamination; and independence from equipment such as ultracentrifuge and chromatography.