Purification and characterization of a β–glucan binding protein from the haemolymph of freshwater prawn Macrobrachium rosenbergii

β-glucan binding protein (βGBP), a pattern recognition protein was purified from the haemolymph of freshwater prawn Macrobrachium rosenbergii by heparin affinity chromatography that showed a single band in native gradient PAGE. The β-glucan binding property of the purified protein was confirmed in a phenoloxidase (PO) assay, where addition of βGBP along with β-glucan increased the specific PO activity compared with that of β-glucan alone. The molecular weight of the βGBP was found to be ~316 kDa on gel filtration chromatography. In SDS-PAGE, βGBP molecule was reduced to one polypeptide chain of molecular weight ~113 kDa. Thus the βGBP in M. rosenbergii is possibly a homotrimeric molecule. The purified sample run on unreduced condition in SDS-PAGE also revealed a similar size band (~113 kDa) and hence, the polypeptide chains of βGBP are held by non-covalent interactions. The purified βGBP samples run in native PAGE was stained positively with alcian blue for carbohydrates and Sudan black for lipids indicating the βGBP to be a glycolipoprotein. With rabbit polyclonal anti-βGBP serum developed, an indirect ELISA was standardized and the normal βGBP concentration in adult M. rosenbergii serum was quantified to be ~2 mg mL−1. Furthermore, the applicability of the developed ELISA is discussed.