Flow cytometric analysis of circulating platelet-monocyte aggregates in whole blood: methodological considerations.

Author(s): Harding SA, Din JN, Sarma J, Jessop A, Weatherall M, Fox KA, Newby DE

Publication: Thromb Haemost, 2007, Vol. 98, Page 451-6

PubMed ID: 17721630 PubMed Review Paper? No

Purpose of Paper

Conclusion of Paper

Studies

1. Study Purpose

   The purpose of this study was to determine the effects of type of anticoagulant, collection method, processing delays, hemolysis, and refrigerated storage of fixed specimens on platelet-monocyte aggregate evaluation by flow cytometry. All experiments were performed with blood from 8 healthy volunteers.

   Summary of Findings:

   Significantly different percentages of platelet-monocyte aggregates were observed between blood specimens incubated for 5 minutes in the different anticoagulants. The highest percentage of aggregates was measured in lithium heparin blood, followed by PPACK, then citrated blood, and finally EDTA blood. Specimens collected via intravenous catheter rather than venipuncture had significantly higher baseline levels of platelet-monocyte aggregation, and platelet-monocyte aggregation increased with sequential collection by intravenous catheter over time, but it did not increase with multiple venipunctures. Room temperature processing delays between 10 and 60 minutes caused platelet-monocyte aggregate percentages to increase for both PPACK and citrated blood (other anticoagulants not tested). Erythrocyte lysis by the addition of FACS lyse solution had no effect on platelet-monocyte aggregation. Storing fixed specimens at 4 degrees C for up to 24 h before analysis had no effects on the percentage of platelet-monocyte aggregates measured.

   Biospecimens

   • Bodily Fluid - Blood

   Preservative Types

   • None (Fresh)
   • Other Preservative

   Diagnoses:

   • Normal

   Platform:

   Analyte	Technology Platform
   Cell count/volume	Flow cytometry

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Biospecimen Acquisition	Anticoagulant	Lithium heparin EDTA PPACK Sodium citrate
   Biospecimen Acquisition	Method of fluid acquisition	IV catheter Venipuncture
   Biospecimen Acquisition	Time of biospecimen collection	Baseline 30 min 50 min 85 min
   Biospecimen Aliquots and Components	Aliquot sequential collection	1st collection 2nd collection 3rd collection 4th collection
   Storage	Time at room temperature	0 min 10 min 20 min 30 min 60 min
   Biospecimen Aliquots and Components	Hemolysis	Chemically-induced Not induced
   Storage	Storage duration	0 h 4 h 8 h 12 h 24 h
   Biospecimen Preservation	Type of fixation/preservation	None (fresh) Refrigeration

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