Anti fcn1

MASP2, FNC1, and C4BPB levels were determined using an in-house ELISA setup. Briefly, polyclonal antibody anti-MASP2, anti-FCN1, or anti-C4BPB (LifeSpan Biosciences) was used to coat overnight at 4 °C 96-well MaxiSorp Nunc plates (Thermo Fisher Scientific, Waltham, MA, USA). The plates were then blocked with 3% BSA in PBS. Exosome fractions were solubilized in 10 µl of mild detergent solution (1% Nonidet P-40, 0.5% Tween-20 in PBS), supplemented with 90 µl 3% BSA in PBST and incubated at 4 °C overnight. After three washes in PBST, the plates were incubated for 4h with the corresponding monoclonal antibody (anti-MASP2, anti-FCN1, or anti-C4BPB, all from LifeSpan Biosciences, Seattle, WA, USA) diluted 1:1000 with 1% BSA in PBST. After three washes with PBST, the plates were incubated with HRP-conjugated anti-mouse IgG diluted 1:5000 in 1% BSA in PBST for 1h. After washing, the peroxidase substrate (TMB, Bio-Rad, Hercules, CA, USA) was added. The reaction was stopped with an H₂SO₄ solution. The absorbance at 450 nm was measured using an iMark microplate reader (Bio-Rad, Hercules, CA, USA). To standardize the response of the antibodies, we used a pool of strongly positive controls. The optical density results were expressed as relative units per milliliter (RU/ml).