Purification and functional analysis of oligomeric OnMBL. (A) The protein concentration of OnMBL in tilapia serum after S. agalactiae challenge was determined by indirect ELISA (n = 4). (B) The expression of OnMBL was analyzed by Western blot at 72 h after bacterial infection. (C) SDS-PAGE (silver stain) analysis of purified serum OnMBL. (D) The b/y ions of the representative peptide of purified protein were analyzed by LC-mass spectrometry/mass spectrometry. (E) Western blot analysis of purified serum OnMBL. (F) Illustration of tilapia MBL molecular weight. (G and H) The oligomers of purified OnMBL from serum of different treatments were analyzed. (I–K) S. agalactiae, LTA, LPS, and mannan binding of oligomeric OnMBL protein (n = 6). (L and M) Oligomeric OnMBL promotes phagocytosis in vitro. Flow cytometric analysis of the MФ phagocytosing S. agalactiae. Data show analyses of 10,000 events. (L) The histogram of flow cytometric analyses of the MФ phagocytosing S. agalactiae preincubated with TBS, BSA, or oligomeric OnMBL (OnMBL^lo and OnMBL^hi). (M) The phagocytic percentage and MFI of MФ, n = 4. (N) Oligomeric OnMBL enhances respiratory burst in MФ. The histogram of flow cytometric analyses of the MФ respiratory burst preincubated with TBS, BSA, OnMBL^lo, or OnMBL^hi protein (10 μg/mL), n = 4.