Two-stage revision arthroplasty is the treatment of choice for periprosthetic infection, a serious complication after knee or hip arthroplasty. Our prospective clinical trial aimed to investigate the concentrations of gentamicin and vancomycin in wound exudate and tissue in two-stage revision arthroplasty. Wound exudate and periprosthetic membrane samples were collected from 18 patients (10 hip and eight knee patients), who were due for two-stage treatment after a periprosthetic joint infection. Samples were taken during insertion of antibiotic-impregnated spacers and after their removal. The concentrations of gentamicin and vancomycin in wound exudates and adjacent tissue were analyzed using high-performance liquid chromatography mass spectrometry. Average time period of spacer implantation was 13.6 weeks (9.3-22.6 weeks). The concentration of vancomycin in wound exudate decreased from a median of 43.28 μg/mL (0.28-261.22) after implantation to 0.46 μg/mL (0.13-37.47) after the removal of the spacer. In the adjacent tissue, vancomycin concentration was mainly undetectable prior to spacer implantation (0.003 μg/g [0.003-0.261]) and increased to 0.318 μg/g [0.024-484.16] at the time of spacer removal. This was also observed for gentamicin in the tissue of patients who previously had cement-free implants (0.008 μg/g [0.008-0.087] vs. 0.164 μg/g [0.048-71.75]) while in the tissue of patients with previously cemented prosthesis, baseline concentration was already high (8.451 μg/g [0.152-42.926]). Despite the rapid decrease in antibiotics release from spacer cement observed in vitro, in vivo antibiotics are much longer detectable, especially in the adjacent soft tissue. © 2018 The Authors. Journal of Biomedical Materials Research Part B: Applied Biomaterials Published By Wiley Periodicals, Inc. J Biomed Mater Res B Part B, 2019. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 107B: 1587-1597, 2019.

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