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首页> 外文期刊>Platelets >Paraformaldehyde fixation induces a systematic activation of platelets.
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Paraformaldehyde fixation induces a systematic activation of platelets.

机译：多聚甲醛固定会诱导血小板的系统活化。

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In whole blood flow cytometric platelet assays sample fixation using paraformaldehyde (PFA) is considered very advantageous to prevent spontaneous activation of platelets in vitro. However, fixation is an important variable in activation assays and its influence on platelets is poorly understood. Using a direct immunofluorescence labelling technique and whole blood flow cytometry, the effect of PFA fixation was investigated for 4 different epitopes on platelet surface each of which mirrors a different aspect of platelet activation, namely P-selectin (CD62P), GP IIbIIa complex (CD41), the fibrinogen binding site of the activated GP IIaIIIb complex (PAC-1) and GP Ib-V-IX complex (CD42b). Platelets fixed with PFA (0.5%) before antibody labelling showed significant (P<0.01) increases in mean fluorescence intensity (MFI) of CD62P (1.10 +/- 0.14 vs. 0.94 +/- 0.12 arbitrary units of fluorescence), CD41 (27.3 +/- 6.3 vs. 15.6 +/- 2.1) and PAC-1 (6.21 +/- 1.25 vs. 0.55 +/- 0.12) when compared to unfixed samples. At the same time, MFI of CD42b was reduced from 28.2 +/- 1.6 to 22.6 +/- 2.3 (P<0.01). When fixation was initiated after antibody labelling, we observed less prominent increases in MFI of CD41 (P<0.05) and PAC-1 (P<0.05) while there was no significant difference for CD62P and rather a moderate rise in CD42b than a decrease (P<0.05). Because these alterations cannot be explained by unspecific effects only, it must be concluded that PFA induces a systematic stimulation of platelets. The lowest in vitro platelet activation was found when antibody labelling was started immediately after blood sampling and when samples were analysed within 10 minutes after being stored without fixation of 4 degrees C in the dark.
机译：在全血流式细胞仪血小板测定中，使用多聚甲醛（PFA）固定样品被认为非常有利于防止体外血小板的自发活化。然而，固定是活化测定中的重要变量，并且其对血小板的影响了解得很少。使用直接免疫荧光标记技术和全血流式细胞术，研究了血小板表面上4个不同表位的PFA固定作用，每个表位反映了血小板活化的不同方面，即P-选择蛋白（CD62P），GP IIbIIa复合物（CD41 ），激活的GP IIaIIIb复合物（PAC-1）和GP Ib-V-IX复合物（CD42b）的纤维蛋白原结合位点。在抗体标记之前用PFA（0.5％）固定的血小板显示CD62P（1.10 +/- 0.14对0.94 +/- 0.12任意荧光单位）的平均荧光强度（MFI）显着（P <0.01）增加与未固定样品相比，+ /-6.3对15.6 +/- 2.1）和PAC-1（6.21 +/- 1.25对0.55 +/- 0.12）。同时，CD42b的MFI从28.2 +/- 1.6降低到22.6 +/- 2.3（P <0.01）。抗体标记后开始固定后，我们观察到CD41和PAC-1的MFI显着上升幅度较小（P <0.05），而CD62P则没有显着差异，而CD42b的上升幅度适中，下降幅度较小（P <0.05）。 P ＜0.05）。因为这些改变不能仅通过非特异性作用来解释，所以必须得出结论，PFA会诱导系统性地刺激血小板。血液采样后立即开始抗体标记，以及在黑暗中未固定4°C保存后10分钟内分析样品时，发现最低的体外血小板活化。

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