In:
BioTechniques, Future Science Ltd, Vol. 54, No. 5 ( 2013-05), p. 271-277
Abstract:
Understanding the response of cells to multiple stimuli is vital for predicting donor specific responses and better understanding the signaling pathways involved. This is of particular importance in platelets because exposure of phosphatidylserine (PS) occurs upon costimulation but not with a single agonist. Here, we describe a multiplexed pairwise agonist scanning-flow cytometry (PAS-FC) method of measuring platelet inside-out responses to all pairs of six platelet agonists (convulxin, SFLLRN, AYPGKF, ADP, U46619, and PGE 2 ) used at their EC 50 concentrations. These agonists allowed exploration of platelet signaling downstream of GPVI, PAR-1, PAR-4, P2Y 1 , P2Y 12 , TP, and IP receptors. The three-color flow cytometry method simultaneously measured integrin α IIb β 3 activation with PAC-1 antibody, P-selectin exposure (via α granule release) with anti-P-selectin, and PS exposure with annexin V. These responses were consistent across a healthy male donor pool. In duplicate measurements with each donor, 4 of the 10 donors had a sufficiently unique 45-parameter (15 pairs × 3 colors) phenotype to self-cluster (P 〈 0.001). This method has the potential for efficiently scanning for patient specific responses across a broad agonist-receptor space.
Type of Medium:
Online Resource
ISSN:
0736-6205
,
1940-9818
Language:
English
Publisher:
Future Science Ltd
Publication Date:
2013
detail.hit.zdb_id:
1496354-1
SSG:
12
