Tobacco is possibly the most promising plant for plant-made pharmaceuticals, e.g., antibodies, due to its high biomass yields and robust transformation technology. Protein A affinity chromatography is commonly used for antibody purification, however, direct application of tobacco extract to protein A chromatography columns may be problematic due to the nonspecific binding of native tobacco proteins (NTPs). Three different protein A resins, ProSep-vA High Capacity, Ultra, and Ultra Plus, from Millipore were studied for antibody purification from tobacco. Experiments were run with extracts from nontransgenic tobacco and the model antibody, human immunoglobulin G (IgG). The efficiency of the wash buffers to reduce nonspecific binding of NTPs to the resins was evaluated by altering the ionic strength and pH. Considering all factors, Ultra Plus Protein A resin may be the best in antibody purification from tobacco.