Monoclonal Antibody Production from Hybridoma Cells

Lc-6 and Lc-4 hybridoma cell cultures [38 (link),39 (link),40 (link),41 (link)] secreting ani-chicken CD4 and CD8, respectively, were maintained in Dulbecco’s Modified Eagle’s Medium (DMEM), as previously described [42 (link)]. The hybridoma cells were initially maintained in DMEM with 20% fetal bovine serum (FBS) that was gradually reduced to 10%, 5%, and 2% (Thermo Fisher Scientific, Waltham, MA, USA) every three days. For antibody collection, cells were maintained in protein-free hybridoma medium II (PFHMII) with 0.2% chemically defined lipid concentrate (Life Technologies, Carlsbad, CA, USA). For large scale mAb production, the Wheaton CELLine 1000 bioreactor flask cells were used following the manufacturer’s protocols (Wheaton, IL, USA).