BACKGROUND: We have previously shown that fresh-frozen plasma (FFP) contains red blood cell-derived procoagulant microparticles (MPs) that are removable by 0.2 microm filtration. Given the limitations of current methods for accurately sizing MPs, we have applied the novel approach of dynamic light scattering (DLS) to characterize the size distributions of these MPs within FFP. METHODS: Fresh-frozen plasma was prepared from blood Group A and O donations (n = 10 of each) after an overnight hold of whole blood at 4 degrees C. On the day of analysis, plasma was thawed to 37 degrees C and daughter aliquots were studied pre- and post-filtration (0.2 microm filtration device, Ceveron MFU-500, Technoclone). MP size and dispersity was assessed using a Zetasizer Nano S (Malvern Instruments Ltd), which employs a 173 degrees backscatter detector and an N5 Submicron Particle Size Analyser (Beckman Coulter) using multi-angle measurements (30.1 degrees , 62.6 degrees and 90 degrees ). The analysers presented MP size distribution graphically as intensity plots, mean size, standard deviation and polydispersity index. RESULTS: Of the instruments used, only the N5 utilizing a 30.1 degrees angle of measurement could detect MPs of the expected size distribution and demonstrate their removal by filtration. MPs (range of mean particle diameters: pre, 101-464 nm; post, 21-182 nm filtration) were significantly smaller post-filtration (P < 0.0001), but polydispersity index (median: pre, 0.746, post, 0.769) exhibited no significant change. There was no significant difference between the size of MPs from blood Group O (pre, 247 nm) and Group A (pre, 289 nm) samples (P = 0.44). CONCLUSION: Our data demonstrates that DLS offers a novel approach to assessing MP size and distribution, a technique that could be easily adopted as a means of assessing MPs within either FFP or other blood products.
BACKGROUND: We have previously shown that fresh-frozen plasma (FFP) contains red blood cell-derived procoagulant microparticles (MPs) that are removable by 0.2 microm filtration. Given the limitations of current methods for accurately sizing MPs, we have applied the novel approach of dynamic light scattering (DLS) to characterize the size distributions of these MPs within FFP. METHODS: Fresh-frozen plasma was prepared from blood Group A and O donations (n = 10 of each) after an overnight hold of whole blood at 4 degrees C. On the day of analysis, plasma was thawed to 37 degrees C and daughter aliquots were studied pre- and post-filtration (0.2 microm filtration device, Ceveron MFU-500, Technoclone). MP size and dispersity was assessed using a Zetasizer Nano S (Malvern Instruments Ltd), which employs a 173 degrees backscatter detector and an N5 Submicron Particle Size Analyser (Beckman Coulter) using multi-angle measurements (30.1 degrees , 62.6 degrees and 90 degrees ). The analysers presented MP size distribution graphically as intensity plots, mean size, standard deviation and polydispersity index. RESULTS: Of the instruments used, only the N5 utilizing a 30.1 degrees angle of measurement could detect MPs of the expected size distribution and demonstrate their removal by filtration. MPs (range of mean particle diameters: pre, 101-464 nm; post, 21-182 nm filtration) were significantly smaller post-filtration (P < 0.0001), but polydispersity index (median: pre, 0.746, post, 0.769) exhibited no significant change. There was no significant difference between the size of MPs from blood Group O (pre, 247 nm) and Group A (pre, 289 nm) samples (P = 0.44). CONCLUSION: Our data demonstrates that DLS offers a novel approach to assessing MP size and distribution, a technique that could be easily adopted as a means of assessing MPs within either FFP or other blood products.
Authors: Heather C Inglis; Ali Danesh; Avani Shah; Jacques Lacroix; Philip C Spinella; Philip J Norris Journal: Cytometry A Date: 2015-04-02 Impact factor: 4.355