Conditions in blood sampling procedures that extend the ex vivo stability of eosinophil activity markers in peripheral blood from allergic patients and healthy controls

Background: Serum-ECP, EG2-epitope on intracellular ECP and surface express ion of CD9 and CD11b in peripheral blood eosinophils (PBE) are considered t o be markers that mirror clinical parameters in allergic inflammation. Objective: The aim was to investigate the impact of the blood sampling proc edure on PBE markers and to identify optimal conditions for extended pre-an alysis storage. Methods: Blood, from healthy individuals and patients with allergic rhiniti s/ asthma, was collected in tubes with EDTA, citrate, or without anti-coagu lant. The expression of EG2-epitope, CD9, and CD11b were analyzed in eosino phils and neutrophils after 1, 5, and 24 hours of storage at +4 degrees C, according to the FOG-method and flow cytometry. In vitro stimulation with f MLP/PMA was used for metabolic activity analysis and CD11b mobilization. Fo llowing a 1-hour clotting period at +20 to 22 degrees C, samples were store d at +4 degrees C and serum-ECP levels were measured. Results: The EG2-epitope, serum-ECP, and CD9 were stable in samples from bo th healthy controls and allergic patients at all storage conditions. The EG 2-epitope, serum-ECP and PBE count were significantly increased in the pati ent group, whereas no differences were observed in the expression of CD9 or CD11b. Both granulocytes and monocytes retained their metabolic activity f or 24 hours. Neutrophils in citrate-blood increased their ability to respon d to fMLP, as compared with EDTA-blood. Conclusion: In vitro analysis of selected activity markers and functional t ests could be performed on granulocytes from both healthy individuals and a llergic patients after 24 hours storage at +4 degrees C. The anticoagulant citrate seems to be preferable to EDTA when monocytes or CD11b expression a re analyzed.