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Isolation and activation of human neutrophils in vitro. The importance of the anticoagulant used during blood collection

Title
Isolation and activation of human neutrophils in vitro. The importance of the anticoagulant used during blood collection
Type
Article in International Scientific Journal
Year
2008
Authors
Marisa Freltas
(Author)
Other
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Graca Porto
(Author)
ICBAS
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Eduarda Femandes
(Author)
FFUP
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Journal
Title: Clinical BiochemistryImported from Authenticus Search for Journal Publications
Vol. 41
Pages: 570-575
ISSN: 0009-9120
Publisher: Elsevier
Scientific classification
FOS: Engineering and technology > Medical engineering
Other information
Authenticus ID: P-003-ZJW
Abstract (EN): Objectives: To assess the effect of different anticoagulants (EDTA, citrate and heparin) on the isolation procedure of human neutrophils and in the subsequent alterations of calcium levels and respiratory burst induced by phorbol myristate acetate (PMA). Design and methods: Isolation of human neutrophils from whole blood was performed by the gradient density centrifugation method. PMA-induced neutrophil burst was measured by chemiluminescence. Intracellular calcium ([Ca2+](i)) was measured using Fluo-3 AM, a calcium-sensitive dye. Results: EDTA provided the highest number of isolated neutrophils/mL of blood (1.7 x 10(6)+/- 1.5 x 10(5)) when compared with citrate (0.46 x 10(6)+/- 0.95 x 10(5)) and heparin (0.66 x 10(6)+/- 0.15 x 10(5)). EDTA originated less degree of PMA-induced activation (370 +/- 30%) relatively to citrate (830 +/- 98%) and heparin (827 +/- 77%). [Ca2+](i) was lower with EDTA (122 +/- 11 nM) when compared with citrate and heparin (150 +/- 13 and 230 +/- 30 nM). Conclusion: The anticoagulant used during blood collection interfered differently with the yield of isolated neutrophils as well as on their calcium levels and reactivity to PMA.
Language: English
Type (Professor's evaluation): Scientific
Contact: earacas@ff.up.pt
No. of pages: 6
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