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Cat. No.: AFAG-200 (for 20-200T)

Cat. No.: AFAG-1k (for 100-1000T)

Description

Autophagosome Fluorescence Staining Kit with AP-Green is a kit for rapid and convenient detection of autophagosomes in live cells using AP-Green as the fluorescent probe. This kit is compatible with fluorescence detection systems including fluorescence microscopes, laser scanning confocal microscopes, flow cytometers, and fluorescence microplate readers. Its performance is highly similar to that of DAPGreen.

Autophagy is an evolutionarily highly conserved intracellular catabolic pathway, which is achieved by lysosomes engulfing and degrading portions of cellular components. Autophagy is closely associated with multiple physiological functions. For instance, under stress conditions such as starvation, cells can degrade excess or abnormal intracellular components via autophagy to supply energy and raw materials for cell survival, thereby sustaining organismal growth and development, cell differentiation, and environmental adaptability. Once autophagy is initiated, cells form a double-membrane isolation membrane that gradually extends to enclose damaged cellular components (such as proteins or organelles), ultimately generating autophagosomes. Autophagosomes then fuse with lysosomes to form autolysosomes. The interior of this structure undergoes gradual acidification to activate lysosomal hydrolases, which further degrade the encapsulated contents. Dysfunctional autophagy is tightly linked to a variety of pathological processes including tumors, neurodegenerative diseases, metabolic disorders, and pathogen infections. Given the critical role of cellular autophagy in physiological and pathological processes, it has become a research hotspot in cell biology.

AP-Green (Autophagosome Green) is a small-molecule lipophilic fluorescent probe with excellent cell membrane permeability. After entering cells, due to its unique structural properties, AP-Green preferentially inserts into the double membrane of autophagosomes during autophagosome biogenesis and accumulates progressively within autophagosomes, emitting enhanced green fluorescence in hydrophobic environments. Leveraging this property, AP-Green enables specific detection of cellular autophagosomes and serves as an important tool for monitoring autophagy. In addition, we offer the Autolysosome Fluorescence Staining Kit (AL-Green) for monitoring autophagosome-lysosome fusion and subsequent degradation.

Based on the staining principle of AP-Green, this kit is only intended for fluorescence staining and detection of autophagosomes in live cells. It cannot be used for staining frozen or fixed cells, tissues, or tissue sections.

Following staining with this kit, imaging can be performed using a fluorescence microscope or confocal microscope, and fluorescence measurement can also be conducted via flow cytometry or fluorescence microplate reader. For fluorescence microscopy, blue excitation light is used to elicit green fluorescence. For confocal microscopy, the recommended excitation wavelength is 450 nm (range: 425–475 nm) and emission wavelength is 535 nm (range: 500–560 nm). For flow cytometry, the FITC channel (Ex/Em = 488/525 nm) is recommended for detection. For fluorescence microplate readers, Ex/Em of 450/535 nm is recommended.

AutophagyUp (1000×) supplied in this kit can be used as a positive control to induce cellular autophagy. The working concentration is generally 0.1–5×, with 1× as the preferred recommended final concentration.

Storage Conditions

Store at -20 ºC, valid for 1 year. AP-Green (1000X) must be protected from light.

Precautions

• AP-Green is extremely light-sensitive. Minimize light exposure during handling.
• Add AP-Green prior to autophagy inducers or inhibitors to enable real-time monitoring of autophagosome formation.
• This kit is for live cell detection only. It is not suitable for fixed or frozen cell or tissue samples.
• This product is for research use only by professionals. It shall not be used for clinical diagnosis or therapy, food or pharmaceutical applications, and must not be stored in residential premises.
• For your safety and health, wear a lab coat and disposable gloves during operation.