Johannes von Behren (Greifswald/ DE), Jan Wesche (Greifswald/ DE), Prof. Dr. Andreas Greinacher (Greifswald/ DE), Dr. Konstanze Aurich (Greifswald/ DE)
Background
Before being implemented in daily clinical routine, new production strategies for platelet concentrates (PC) must be validated for their safety. Besides in vitro testing, the establishment of new methods requires the labeling of platelets for in vivo studies of platelets" survival and recovery. Indocyanine green (ICG) is a FDA and BfArM approved near-infrared (NIR) fluorescent dye suitable for non-radioactive direct labeling of platelets for diagnostics.
Methods
Platelets from PC in storage solutions with different plasma concentrations were labeled with ICG up to concentrations of 200 µM. Whole blood was used as an ex vivo matrix to monitor the labeling stability of ICG-labeled platelets. The impact of labeling processes was assessed by the quantification of CD62P expression and PAC-1 binding as platelet function markers. Platelet aggregation was analyzed by light transmission aggregometry. ICG labeling efficiency and stability of platelets was determined by flow cytometry.
Results
Platelets from PC were successfully labeled with 10 µM ICG after 1 and 4 days of storage. The best labeling efficiency of 99.8% ± 0.1% (immediately after labeling) and 81% ± 6.2% (after 24 hours incubation with whole blood) was achieved by plasma replacement with 100% platelet additive solution for the labeling process. The washing process slightly impaired platelet function, but ICG labeling itself did not affect platelets. Immediately after the ICG labeling process, plasma was re-added resulting in a recovered platelet function.
Conclusion
We developed a Good Manufacturing Practice compatible protocol for indocyanine fluorescent platelet labeling suitable for non-radioactive tracing in survival and recovery studies in vivo.
Offenlegung Interessenkonflikt:
none