Formaldehyde-fixation of platelets for flow cytometric measurement of phosphatidylserine exposure is feasible.
Details
Serval ID
serval:BIB_8E89A4C92EF3
Type
Article: article from journal or magazin.
Collection
Publications
Institution
Title
Formaldehyde-fixation of platelets for flow cytometric measurement of phosphatidylserine exposure is feasible.
Journal
Cytometry. Part A
ISSN
1552-4930 (Electronic)
ISSN-L
1552-4922
Publication state
Published
Issued date
2015
Peer-reviewed
Oui
Volume
87
Number
1
Pages
32-36
Language
english
Notes
Publication types: Journal Article
Publication Status: ppublish
Publication Status: ppublish
Abstract
Strong platelet activation results in a redistribution of negatively charged phospholipids from the cytosolic to the outer leaflet of the cellular membrane. Annexin V has a high affinity to negatively charged phospholipids and can be used to identify procoagulant platelets. Formaldehyde fixation can cause factitious Annexin V binding. Our aim was to evaluate a method for fixing platelets avoiding additional Annexin V binding. We induced expression of negatively charged phospholipids on the surface of a fraction of platelets by combined activation with convulxin and thrombin in the presence of Annexin V-fluorescein isothiocyanate and calcium. Aliquots of resting and activated platelets were fixed with a low concentration, calcium-free formaldehyde solution. Both native platelets and fixed platelets were analyzed by flow cytometry immediately and after a 24-h storage at 4°C. We observed that the percentage of Annexin V positive resting platelets ranged from 1.5 to 9.3% for the native samples and from 0.4 to 12.8% for the fixed samples (P=0.706, paired t-test). The amount of Annexin V positive convulxin/thrombin activated platelets varied from 12.9 to 35.4% without fixation and from 15.3 to 36.3% after formalin fixation (P=0.450). After a 24-h storage at 4°C, Annexin V positive platelets significantly increased both in the resting and in the convulxin/thrombin activated samples of native platelets (both P<0.001), while results for formalin fixed platelets did not differ from baseline values (P=0.318 for resting fixed platelets; P=0.673 for activated fixed platelets). We conclude that platelet fixation with a low concentration, calcium-free formaldehyde solution does not alter the proportion of Annexin V positive platelets. This method can be used to investigate properties of procoagulant platelets by multicolor flow-cytometric analysis requiring fixation steps.
Pubmed
Web of science
Open Access
Yes
Create date
10/02/2015 9:21
Last modification date
20/08/2019 14:52