Utilizing lock-in amplification to measure vital parameters during ex situ liver perfusion

Abstract Liver transplantation remains the only treatment for end-stage liver disease; however, its application is critically constrained by the persistent shortage of viable donor organs. This shortage motivates the use of extended criteria donor livers, which increases the risk for primary non-function. Accurately assessing graft quality of marginal grafts remains a challenge due to a lack of predictive markers. Ex situ liver perfusion (EVLP) has emerged as a potential strategy to increase the time for assessment by introducing a platform that increases preservation times and mimics the in vivo environment for the graft. Unlike for static cold storage (SCS), oxygenated perfusates are continuously pumped through donor livers during EVLP providing an opportunity for viability assessment and potential rehabilitation of marginal grafts prior to transplantation. The transition from passive preservation via SCS to active EVLP enables continuous access to the perfusate of the metabolically active graft, enabling automatic real-time monitoring of biomarkers. Here, we describe the design and application of a real-time optical setup based on lock-in amplification for continuous monitoring of critical liver viability biomarkers–indocyanine green (ICG) and flavin mononucleotide (FMN)–during EVLP. ICG is well-established to visualize hepatic blood flow and clearance correlates with hepatic function. ICG is measured directly within the blood-filled perfusion tubing. This approach leverages the near-infrared absorbance of ICG to mitigate optical interference arising from blood turbidity. Concurrently, FMN, an early marker of mitochondrial damage and ischemia–reperfusion injury (IRI), is optically quantified in the transparent perfusate during hypothermic oxygenated machine perfusion or in waste dialysis during normothermic machine perfusion, effectively bypassing the optical interference inherent in direct blood measurements. This optical design successfully enabled continuous, quantitative measurements of both ICG and FMN, providing dynamic insights into IRI and metabolic status of ex situ perfused human and rodent livers, paving the way for further assessment of donor livers.