Marine Ma , Staining fixed cells is not recommended. The principle of ER-Tracker Green staining typically relies on the physiological activity of cells or the specific structural/functional state of the endoplasmic reticulum. However, fixatives can crosslink intracellular proteins and disrupt the natural structure of the endoplasmic reticulum (ER), such as membrane integrity and the luminal microenvironment. This can directly disrupt the dye's binding sites to the ER. This can lead to cell viability after fixation and an inability to maintain the physiological conditions required for targeted dye binding.

Staining before fixation is an option, but the fluorescence signal may be partially retained. Be aware that aldehyde fixation (such as 4% paraformaldehyde) may quench or weaken some fluorescence signals. It is recommended to verify the preservation of fluorescence signals and structural integrity in advance, and to include negative controls (such as unstained fixation) to eliminate interference.

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