巨噬细胞和嗜中性粒细胞等细胞可以内化一组不同的颗粒物,说明细菌和凋亡小体经由过程吞噬。这些颗粒被隔离成吞噬体,然后与早期和晚期内体融合,最终与溶酶体融合成熟成吞噬溶酶体,通过一个被称为吞噬体成熟的过程。最终,粒子降解后,然后,吞噬体片段以通过吞噬体分解来改革溶酶体。随着吞噬体的变化,它们获取和剥离与吞噬体成熟和分解的各个阶段相关的蛋白质。可以通过使用免疫荧光方法在单吞噬体水平上评估这些变化。通常,我们使用间接免疫荧光方法,该方法依赖于针对追踪吞噬体成熟的特定分子标记的一级抗体。通常,吞噬小体进入吞噬溶酶体的进展可以通过对细胞进行溶酶体相关膜蛋白I(LAMP1)染色并通过显微镜或流式细胞术测量每个吞噬小体周围LAMP1的荧光强度来确定。然而,该方法可用于检测任何具有免疫荧光相容抗体的分子标记。
Cells such as macrophages and neutrophils can internalize a diverse set of particulate matter, illustrated by bacteria and apoptotic bodies through the process of phagocytosis. These particles are sequestered into
phagosomes, which then fuse with early and late endosomes and ultimately with lysosomes to mature into phagolysosomes, through a process known as phagosome maturation. Ultimately, after particle degradation,
phagosomes then fragment to reform lysosomes through phagosome resolution. As
phagosomes change, they acquire and divest proteins that are associated with the various stages of phagosome maturation and resolution. These changes can be assessed at the single-phagosome level by using immunofluorescence methods. Typically, we use indirect immunofluorescence methods that rely on primary antibodies against specific molecular markers that track phagosome maturation. Commonly, progression of
phagosomes into phagolysosomes can be determined by staining cells for Lysosomal-Associated Membrane Protein I (LAMP1) and measuring the fluorescence intensity of LAMP1 around each phagosome by microscopy or flow cytometry. However, this method can be used to detect any molecular marker for which there are compatible antibodies for immunofluorescence.