Figure 1.

1a) HPLC profile of BTL-2 using an ion exchange column, protein Pack SP 5PW (Waters). Approximately 5 mg of the sample, obtained from the DEAE column, were dissolved in 0.05 M ammonium bicarbonate and applied onto the chromatographic column. The chromatography showed the presence of two main fractions containing haemagglutinating activity, named BTL-1 and BTL-2. The run was monitored at 280 nm and the elution was conducted using a discontinuous linear gradient of ammonium bicarbonate at a concentration of 1.0 M, pH 8.0, constant flow rate of 1 mL/min. 1b) The BTL-2 fraction was subjected to a final chromatography in a Reverse phase HPLC fractionation using a μ-Bondapack C18 column. The elution of samples was carried out using a linear gradient concentration of an aqueous solution of 66% acetonitrile and was monitored at 280 nm at a constant flow rate of 1 mL/min. Samples were previously dissolved in buffer A (0.1% TFA) that was used to equilibrate the column. 1c) Following the method [43], a discontinuous electrophoresis was performed with a final acrylamide concentration of 12% in the running gel. All the samples and the molecular markers were treated with SDS and DTT 1 M and the run was conducted at 40 mA. The samples were stained with Coomassie brilliant blue R-250. The SDS-PAGE profile of BTL-2 showed high molecular homogeneity in a single band with estimated molecular mass of approximately 9.0 kDa.