Fluo-4 AM is a commonly used probe for detecting intracellular Ca2+ concentration at a wavelength of 494/506 nm.
Fluo-4 is a calcium fluorescent probe that replaces Cl with F in the Fluo-3 structure. Due to the replacement of Cl with the more electronically attractive F, its excitation wavelength deviates by about 10 nm toward the short wavelength. This wavelength is closer to that of an argon laser, so the fluorescence intensity of Fluo-4 is twice as strong as that of Fluo-3 when excited with an argon laser.
Fluo-4 AM is a cell-permeable fluorescent Ca2+ indicator (Kd Ca2+ = 345 nM). Fluo-4 AM penetrates the cell membrane and enters the cell, then is sheared by intracellular esterase to form Fluo-4, which is retained in the cell. Fluo-4 is almost non-fluorescent when it exists in the form of free ligand, but when it combines with intracellular calcium ions, it can produce strong fluorescence, and the intensity of fluorescence is increased by nearly 100 times.
When heavy metals are present in solution (e.g. Mn2+, Zn2+, Pb2+) , the heavy metal-selective chelator TPEN (Catalog No. GC12918) can be used to control perturbations in calcium measurements caused by these ions. Fluo-4 AM is suitable for fluorescence and confocal microscopy, flow cytometric analysis, and microplate screening applications.
Fluo-4 AM 是一种常用的检测细胞内 Ca2+ 浓度的探针，检测波长为494/506 nm。
Fluo-4是一种将Fluo-3结构中的Cl替换成F的钙荧光探针。由于将Cl替换成了电子吸引力更强的F，它的激发波长会向短波长处偏离10 nm左右。这个波长更接近于氩激光器的波长，所以用氩激光器激发时，Fluo-4的荧光强度比Fluo-3强一倍。
Fluo-4 AM是一种细胞渗透性荧光Ca2+指示剂（Kd Ca2+ = 345 nM）。 Fluo-4 AM穿透细胞膜进入细胞后被细胞内的酯酶剪切形成Fluo-4，从而被滞留在细胞内，Fluo-4若以游离配体形式存在时几乎是非荧光性的，但是当它与细胞内钙离子结合后可以产生较强的荧光，荧光强度增加近100 倍。
当溶液中存在重金属时（例如 Mn2+、Zn2+、Pb2+），可以使用重金属选择性螯合剂 TPEN （目录号：GC12918）来控制由这些离子引起的钙测量值扰动。Fluo-4 AM适用于荧光和共聚焦显微镜、流式细胞分析和微孔板筛选应用。