Fura-4F AM

Operating Instructions
(The following is a recommended experimental protocol for guidance only, and adjustments are required based on your specific needs)

1. Stock Solution Preparation
1.1 Solvent: Most dyes are typically dissolved in organic solvents, such as anhydrous DMSO.
1.2 Concentration Recommendation: It is generally recommended to prepare a high-concentration stock solution of 1-10 mM.

2. Working Solution Preparation
2.1 Diluent: Modified Krebs-Henseleit solution.
2.2 Working Concentration: 2.5 μM.
2.3 Notes: Prepare and use immediately.

3. Staining Procedure
3.1 Sample Types:
3.1.1 Intact heart tissue: Langendorff-perfused intact guinea pig hearts.
3.1.2 Tissue sections: Guinea pig left ventricular tissue sections (350 μm thick).
3.2 Incubation Conditions: Incubate with 2.5 μM fura-4F AM working solution for 30 min at 36±1 °C via cyclic perfusion.
3.3 Washing Step: Information not provided; this section is omitted.

4. Control Group
Information not provided; this section is omitted.

5. Detection and Analysis
5.1 Instrument: EMCCD high-speed camera.
5.1.1 Excitation wavelength: 385 nm; emission wavelength is filtered through a multi-band filter with a passband for calcium dye signals.
5.2 Result Analysis:
5.2.1 Changes in fluorescence intensity: Fura-4F AM can record intracellular Ca²⁺ transient signals in isolated hearts and heart tissue sections. After treatment with Nifedipine, the CaT amplitude decreases, indicating that this dye can be used to monitor drug-induced Ca²⁺ dynamic changes.
5.2.2 This dye is used for Ca²⁺ optical imaging in whole hearts and tissue sections in literature, with no description of subcellular localization.