Staphylococcus epidermidis is a leading cause of biofilm-associated infections on implanted medical devices. During the treatment of an infection, bacterial cells inside biofilms may be exposed to sublethal concentrations of the antimicrobial agents. In the present study, the effect of subinhibitory concentrations of tigecycline (TC) on biofilms formed by S. epidermidis strain RP62A was investigated using a quantitative global proteomic technique. Sublethal concentrations of TC [1/8 (T1) and 1/4 minimum inhibitory concentration (MIC) (T2)] promoted biofilm production in strain RP62A, but 1/2 MIC TC (T3) significantly inhibited biofilm production. Overall, 413, 429, and 518 proteins were differentially expressed in biofilms grown with 1/8 (T1), 1/4 (T2), and 1/2 (T3) MIC of TC, respectively. As the TC concentration increased, the number of induced proteins in each Cluster of Orthologous Groups (COG) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway increased. The TC concentration dependence of the proteome response highlights the diverse mechanisms of adaptive responses in strain RP62A biofilms. In both COG and KEGG functional analyses, most upregulated proteins belong to the metabolism pathway, suggesting that it may play an important role in the defense of strain RP62A biofilm cells against TC stress. Sub-MIC TC treatment of strain RP62A biofilms led to significant changes of protein expression related to biofilm formation, antimicrobial resistance, virulence, quorum sensing, ABC transporters, protein export, purine/pyrimidine biosynthesis, ribosomes, and essential proteins. Interestingly, in addition to tetracycline resistance, proteins involved in resistance of various antibiotics, including aminoglycosides, antimicrobial peptides, ß-lactams, erythromycin, fluoroquinolones, fusidic acid, glycopeptides, lipopeptides, mupirocin, rifampicin and trimethoprim were differentially expressed. Our study demonstrates that global protein expression profiling of biofilm cells to antibiotic pressure may improve our understanding of the mechanisms of antibiotic resistance in biofilms.