Platelet Functions During Extracorporeal Membrane Oxygenation. Platelet–Leukocyte Aggregates Analyzed by Flow Cytometry as a Promising Tool to Monitor Platelet Activation
Abstract
:1. Introduction
2. Platelet Activation, Aggregation, and ECMO
3. Inflammation in ECMO Patients
4. Inflammation and Platelet–Leukocyte Interactions
5. Flow Cytometric Whole Blood Measurement of Platelet–Leukocyte Aggregates
5.1. General Considerations
5.2. Pre-Analytical Requirements—Choice of Anticoagulant, Delay between Sampling and Immunolabeling, Sample Stability after Immunolabeling, Effect of Strong Vortex Agitation prior to Immunolabeling
5.3. Optimal Conditions for Whole Blood Cytometric Measurement of PLAs: A Proposal
6. Conclusions
Author Contributions
Funding
Conflicts of Interest
References
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Type of ECMO and PumpNumber of Patients Study Reference | Blood Collection | Results |
---|---|---|
Centrifugal pump Jostra pumphead, Maquet® VA n = 7; VV n = 3 Nair et al. [12] | During ECMO. Citrated whole blood. | Patients with platelet count > 100 × 109/L 50% to 72% of patients within low range of ADP, TRAP, collagen aggregation, and ristocetin agglutination |
VA n = 26; VV n = 12 Tauber et al. [13] | Before, after 24 h and 48 h initiation of ECMO therapy and 24 h after ECMO termination. Hirudin anticoagulated whole blood. | 30% to 40% decrease in aggregation with ADP, TRAP, AA after 24 h/baseline. After 48 h, aggregation with TRAP = baseline, ADP and AA lower/baseline Return to baseline after 24 h |
ROTAFLOW® or CARDIOHELP® centrifugal pumps. VV n = 20 Wand et al. [14] | Before, 6 h, 1, 2, 3 and 7 days after the start of ECMO therapy. Whole blood sample on heparin-anticoagulated and calcium-balanced tubes. | Patients with platelet count > 70 × 109/L. Reduced platelet aggregation with ADP, TRAP, AA 6 h after ECMO initiation/before, spontaneous recovery on day 2 with values exceeding baseline afterwards |
VA n = 23; VV n = 10 Balle et al. [15] | Every day from day 1 to day 7 in 33 patients. Hirudin anticoagulated whole blood. | Platelet aggregation with ADP, TRAP, AA: lower compared to healthy volunteers from day 1 up to day 7 but similar when analyzed relative to platelet count |
Sample Handling and Preparation | |
---|---|
Anticoagulant in blood sampling tube | Preferably CTAD (other option: sodium citrate) |
Storage temperature | Room temperature |
Centrifugation before immunolabeling | None (whole blood protocol) |
Prefixation before immunolabeling | None |
Erythrocyte lysis before immunolabeling | None |
Processing time before immunolabeling | Fixed time to allow comparative analysis As short as possible, preferably < 1 h after blood withdrawal |
Fixation after immunolabeling | Optional, paraformaldehyde fixation (0.5 to 1%) |
Final blood dilution | ≥1:40 |
Erythrocyte lysis after immunolabeling | Optional |
Processing time after immunolabeling | Without delay if no fixation 24 h at 4 °C, after fixation |
Cytometric flow rates | Low to medium, determined in each laboratory |
Immunolabeling | |
Platelet-specific antibody | Preferably: CD41 Other option: CD61 |
Leukocyte-specific antibody | CD45 |
Leukocyte subtype-specific antibody | Neutrophils: CD66b |
Monocytes: CD14 | |
T-cells: CD3, CD4 and CD8 | |
B-cells: CD19 | |
NK-cells: CD56 |
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Mansour, A.; Roussel, M.; Gaussem, P.; Nédelec-Gac, F.; Pontis, A.; Flécher, E.; Bachelot-Loza, C.; Gouin-Thibault, I. Platelet Functions During Extracorporeal Membrane Oxygenation. Platelet–Leukocyte Aggregates Analyzed by Flow Cytometry as a Promising Tool to Monitor Platelet Activation. J. Clin. Med. 2020, 9, 2361. https://doi.org/10.3390/jcm9082361
Mansour A, Roussel M, Gaussem P, Nédelec-Gac F, Pontis A, Flécher E, Bachelot-Loza C, Gouin-Thibault I. Platelet Functions During Extracorporeal Membrane Oxygenation. Platelet–Leukocyte Aggregates Analyzed by Flow Cytometry as a Promising Tool to Monitor Platelet Activation. Journal of Clinical Medicine. 2020; 9(8):2361. https://doi.org/10.3390/jcm9082361
Chicago/Turabian StyleMansour, Alexandre, Mikael Roussel, Pascale Gaussem, Fabienne Nédelec-Gac, Adeline Pontis, Erwan Flécher, Christilla Bachelot-Loza, and Isabelle Gouin-Thibault. 2020. "Platelet Functions During Extracorporeal Membrane Oxygenation. Platelet–Leukocyte Aggregates Analyzed by Flow Cytometry as a Promising Tool to Monitor Platelet Activation" Journal of Clinical Medicine 9, no. 8: 2361. https://doi.org/10.3390/jcm9082361
APA StyleMansour, A., Roussel, M., Gaussem, P., Nédelec-Gac, F., Pontis, A., Flécher, E., Bachelot-Loza, C., & Gouin-Thibault, I. (2020). Platelet Functions During Extracorporeal Membrane Oxygenation. Platelet–Leukocyte Aggregates Analyzed by Flow Cytometry as a Promising Tool to Monitor Platelet Activation. Journal of Clinical Medicine, 9(8), 2361. https://doi.org/10.3390/jcm9082361