Transdermal drug delivery is an attractive route of administration relative to other routes as it offers enhanced therapeutic efficacy. However, due to poor skin permeability of certain drugs, their application in transdermal delivery is limited. The ultra-deformable nature of transferosomes makes them suitable vehicles for transdermal delivery of drugs that have high molecular weights and hydrophilicity. However, their low viscosity, which leads to low contact time on the surface of the skin, has restricted their application in transdermal delivery. Therefore, this study aimed to deliver transferosomes loaded with a highly water-soluble and high molecular weight vancomycin hydrochloride (VCM-HCl) via a bigel for systemic delivery and treatment of microbial infections. VCM-HCl-loaded transferosomal formulations (TNFs) were prepared using a reverse-phase evaporation method and then loaded into a bigel. Both the TNFs and TNFs-loaded bigel (TNF-L-B) were characterized by a range of in vitro and ex vivo techniques. TNFs and TNF-L-B were tested for biosafety via the MTT assay and found to be biosafe. Prepared TNFs had sizes, zeta potential and entrapment efficiency of 63.02 ± 5.34 nm, -20.93 ± 6.13 mV and 84.48 ± 1.22% respectively. VCM-HCl release from TNF-L-B showed a prolonged release profile with 39.76 ± 1.6% after 24hrs when compared to bare VCM-HCl loaded in the bigel (74.81 ± 8.84%). Ex-vivo permeation of prepared TNF-L-B showed a higher permeation flux of 0.56 µg/cm2/h compared to the bare VCM-HCl-loaded bigel of 0.23 µg/cm2/h, indicating superior permeation and bioavailability of the drug. Additionally, the prepared TNF-L-B demonstrated improved antimicrobial activity. The TNF-L-B showed minimum inhibitory concentrations (MIC) of 0.97 µg/ml against Staphylococcus aureus (SA) and 1.95 µg/ml against methicillin-resistant SA (MRSA), which were 2-fold lower MIC values than the bare drug. The time-kill assay showed that both TNFs and TNF-L-B systems caused a 5.6-log reduction (100%) in MRSA compared to bare VCM-HCl after 24 hrs of incubation. Furthermore, as opposed to the bare VCM-HCl solution, the degree of biofilm reduction caused by TNFs (55.72%) and TNF-L-B (34.58%) suggests their dominance in eradicating MRSA biofilm. These findings indicate that TNF-L-B is a promising system for transdermal delivery of hydrophilic and high molecular weight drugs.