Background: Mesenchymal stem/stromal cells (MSCs) have demonstrated promise in pathogenic acute respiratory distress syndrome models and are advancing to clinical efficacy testing. Besides immunomodulatory effects, MSC derived conditioned medium (CM) has direct antibacterial effects, possibly through LL-37 and related secreted peptide activity. We investigated MSC-CM compatibility with vibrating mesh technology, allowing direct delivery to the infected lung. Methods: MSC-CM from bone marrow (BM) and umbilical cord (UC) MSCs were passed through the commercially available Aerogen Solo nebulizer. Known colony forming units of Escherichia coli, Staphylococcus aureus, and multidrug resistant Klebsiella pneumoniae clinical isolates were added to MSC-CM in an orbital shaker and antibacterial capacity assessed through OD600 spectrophotometry. To exclude the possible effects of medium depletion on bacteria proliferation, MSC-CM was concentrated with a 3000 Da cutoff filter, diluted with fresh media, and retested against inoculum. Enzyme-linked immunosorbent assay was used to quantify levels of antimicrobial peptides (AMPs) and IL-8 present at pre- and postnebulization. Results: Both BM and UC MSC-CM inhibited proliferation of all pathogens, and this ability was retained after nebulization. Concentrating and reconstituting CM did not affect antibacterial properties. Interestingly, LL-37 protein did not appear to survive nebulization, although other secreted AMPs and an unrelated protein, IL-8, were largely intact. Conclusion: MSC-CM is a potent antimicrobial agent and is compatible with vibrating mesh nebulization delivery. The mechanism is through a secreted factor that is over 3000 Da in size, although it does not appear to rely solely on previously identified peptides such as LL-37, hepcidin, or lipocalin-2.