Absolute Quantification of Plasma Membrane Receptors Via Quantitative Flow Cytometry.
Absolute Quantification of Plasma Membrane Receptors Via Quantitative Flow Cytometry.
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DOI：
10.1007/978-1-0716-2217-9_4
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发表时间：
2022
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质膜受体是一种跨膜蛋白，可在特定的配体(如生长因子、激素和细胞因子)结合后启动细胞反应。在许多人类疾病中，质膜受体的丰度可以作为诊断或预后的生物标志物。定量流式细胞术(QFlow)是检测质膜受体最好的技术之一。QFlow使用针对感兴趣受体的荧光团结合抗体，相应的荧光团负载校准珠提供标准化和可重复性的质膜受体测量。更重要的是，当藻红蛋白(PE)作为荧光载体时，qFlow可以实现质膜受体的绝对定量。在这里，我们描述了一个详细的qFlow协议，以获得基于PE校准的绝对受体数量。该方案为许多先前和正在进行的利用体外细胞模型和体外细胞样本量化酪氨酸激酶受体的研究奠定了基础。
Plasma membrane receptors are transmembrane proteins that initiate cellular response following the binding of specific ligands (e.g., growth factors, hormones, and cytokines). The abundance of plasma membrane receptors can be a diagnostic or prognostic biomarker in many human diseases. One of the best techniques for measuring plasma membrane receptors is quantitative flow cytometry (qFlow). qFlow employs fluorophore-conjugated antibodies against the receptors of interest and corresponding fluorophore-loaded calibration beads offers standardized and reproducible measurements of plasma membrane receptors. More importantly, qFlow can achieve absolute quantification of plasma membrane receptors when phycoerythrin (PE) is the fluorophore of choice. Here we describe a detailed qFlow protocol to obtain absolute receptor quantities on the basis of PE calibration. This protocol is foundational for many previous and ongoing studies in quantifying tyrosine kinase receptors with in-vitro cell models and ex-vivo cell samples.