Immunofluorescence Protocol for Characterization of Platelet and Leukocyte Binding in Extracorporeal Membrane Oxygenation (ECMO) Circuits.
Tengyi CaiMatthew BurtonConor McCaffertySuelyn Van Den HelmNatasha LetunicaChantal AttardStephen HortonSteve BottrellBradley SchultzGraeme MacLarenRoberto ChilettiDerek BestAmy JohansenFiona NewallWarwick ButtYves d'UdekemPaul MonagleVera IgnjatovicPublished in: ASAIO journal (American Society for Artificial Internal Organs : 1992) (2024)
The continuous contact between blood and the foreign surface of the extracorporeal membrane oxygenation (ECMO) circuit contributes to hemostatic, inflammatory, and other physiological disturbances observed during ECMO. Although previous studies have extensively investigated blood samples from patients on ECMO, cell adsorption to the ECMO circuit as an additional factor that could potentially influence clinical outcomes, has largely been overlooked. Here we provide a detailed immunofluorescence (IF) protocol designed to characterize cellular binding on ECMO circuits collected from patients. Extracorporeal membrane oxygenation circuits were collected from three pediatric patients and an albumin primed-only ECMO circuit was used as control. Circuit samples from five different sites within each ECMO circuit were collected and processed for the IF protocol. CD14 and CD42a antibodies were used to identify platelets and leukocytes bound to each ECMO circuit sample and images captured using inverted fluorescence microscopy. The protocol enables the comprehensive characterization of platelet and leukocyte binding to ECMO circuits collected from patients, which could in turn extend our knowledge of the characteristics of circuit binding and may provide guidance for improved ECMO circuit design.
Keyphrases
- extracorporeal membrane oxygenation
- acute respiratory distress syndrome
- respiratory failure
- end stage renal disease
- chronic kidney disease
- ejection fraction
- newly diagnosed
- randomized controlled trial
- mechanical ventilation
- peritoneal dialysis
- healthcare
- prognostic factors
- intensive care unit
- single molecule
- peripheral blood
- patient reported outcomes
- high throughput
- stem cells
- high resolution
- bone marrow
- mesenchymal stem cells
- binding protein
- transcription factor
- single cell
- high speed
- case control
- energy transfer