Whole blood was diluted in 0.9 % NaCl solution in a 1:1 ratio and lymphocytes were isolated by density gradient centrifugation using lymphoprep solution (Axis-shield PoC, Oslo, Norway) upon centrifugation at 1800 × g at 20 °C for 20 min. A clear distinct layer of mononuclear cells containing lymphocytes was carefully pipetted from the tube and cultured in L-15 medium (ATCC, Borås, Sweden), supplemented with 10 % fetal bovine serum (FBS; Sigma Aldrich, Stockholm, Sweden), at 37 °C. The lymphocytes were then stimulated with heat-killed E. fecalis (CCUG, Gothenberg University, Sweden) at an optical density (OD) of 1 and incubated at 37 °C for 3 weeks (Additional file 1). After 3 weeks, the medium was collected and centrifuged at 3000 × g for 10 min to pellet the cells. The supernatant was filtered using 0.45 μm sterile filters (Thermofisher Scientific, Stockholm, Sweden) and centrifuged again using 100 KDa Amicon Microcon Centrifugal Filters (Millipore, Molsheim, France) at 3000 × g for 60 min. Antibodies with molecular weight over 100KDa were recovered from the filters by pipetting and were stored at −70 °C until further use.