Lyso id green detection kit

Manufactured by Enzo Life Sciences

Sourced in United States

The Lyso-ID Green detection kit is a fluorescent probe designed to detect and quantify lysosomal activity in cells. The kit provides a sensitive and specific method for measuring lysosomal function.

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Lab products found in correlation

Nucleocounter nc 3000 system, by ChemoMetec (3 mentions) Lsrfortessa, by BD (1 mentions) Glass bottom dishes, by Ibidi (1 mentions)

7 protocols using lyso id green detection kit

Lysosomal Signal Detection in HAoEC

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HAoEC were grown to confluence and treated as indicated. For detection of pH-dependent lysosomal signal, HAoEC were incubated with LysoID Green detection kit (ENZ51034; EnzoLife Sciences, USA) as recommended by the manufacturer and detached from surfaces using the PromoCell split kit. For flow cytometry-based quantification, cells were washed after the incubation and analysed immediately using the LSR Fortessa (BD Bioscience, USA). Data were analysed using FlowJo software (FlowJo9/10, LLC, Ashland, Ore).
For imaging of LysoID, cells were washed after incubation and carefully covered with a glass cover slip and imaged immediately (Echo Revolve Microscope) with the same exposure time throughout each imaging experiment.

Baumer Y., Dey A.K., Gutierrez-Huerta C.A., Khalil N.O., Sekine Y., Sanda G.E., Zhuang J., Saxena A., Stempinski E., Elnabawi Y.A., Dagur P.K., Ng Q., Teague H.L., Keel A., Rodante J.A., Boisvert W.A., Tsoi L.C., Gudjonsson J.E., Bleck C.K., Chen M.Y., Bluemke D.A., Gelfand J.M., Schwartz D.M., Kruth H.S., Powell-Wiley T.M., Playford M.P, & Mehta N.N. (2020). Hyperlipidaemia and IFNgamma/TNFalpha Synergism are associated with cholesterol crystal formation in Endothelial cells partly through modulation of Lysosomal pH and Cholesterol homeostasis. EBioMedicine, 59, 102876.

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Quantifying Acidic Vesicles in Live Cells

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Acidic vesicles were detected in live cells using the Lyso-ID Green detection kit (Enzo Lifesciences, Farmingdale, NY, USA). Cells were seeded in 12 well plates (90% confluent) and incubated with 4 µl/ml Lyso-ID in assay buffer at 37 °C for 30 minutes. Cells were washed with assay buffer and fluorescence measured on a plate reader at excitation 488 nm, emission 520 nm. Following reading, buffer was aspirated, cells lysed overnight in 0.25 M NaOH and protein concentration measured. Cell fluorescence was expressed as fluorescent units/mg protein.

Magalhaes J., Gegg M.E., Migdalska-Richards A, & Schapira A.H. (2018). Effects of ambroxol on the autophagy-lysosome pathway and mitochondria in primary cortical neurons. Scientific Reports, 8, 1385.

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Measuring Lysosome Formation Induced by MG

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Lysosome formation induced by MG was measured using the LYSO-ID® Green Detection Kit (ENZ-51034, Enzo). 2.4×10⁵ cells were seeded into each well of six-well plates and cultured in a CO₂ incubator at 37°C overnight. MG or the vehicle were used to treat the cells for 24 h, and the cells were harvested and stained with fluorescent dyes using the LYSO-ID® Green Detection Kit as described by the manufacturer’s. Fluorescence intensity was measured using the NucleoCounter® NC-3000^TM system (ChemoMetec) [37 (link)].

Huang C.Y., Chang Y.J., Wei P.L., Hung C.S, & Wang W. (2021). Methyl gallate, gallic acid-derived compound, inhibit cell proliferation through increasing ROS production and apoptosis in hepatocellular carcinoma cells. PLoS ONE, 16(3), e0248521.

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Lysosome Detection in Hypoxia

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Cells were plated on 8-well coverglass slide (Sarstedt, Germany Cat.n° 94 6190802) and treated with LPS under normoxic or hypoxic conditions. For Baf A1 and CQ treatment, the compounds were added at a concentration of 100 nM and 100 µM, respectively, 6 h before the end of the experiment. After 24 h, cells were labeled by Lyso-ID Green Detection Kit (Enzo Life Sciences, Plymouth Meeting, PA, USA), and nuclear staining was performed by using DAPI. Cells were analyzed by confocal microscope and the fluorescence intensity was determined by ImageJ software, as described above.

Monaci S., Aldinucci C., Rossi D., Giuntini G., Filippi I., Ulivieri C., Marotta G., Sozzani S., Carraro F, & Naldini A. (2020). Hypoxia Shapes Autophagy in LPS-Activated Dendritic Cells. Frontiers in Immunology, 11, 573646.

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Quantifying ANE-Induced Lysosome Formation

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ANE-induced lysosome formation was measured by a LYSO-ID® Green detection kit (ENZ-51034, Enzo). The dye accumulates in acidic compartments, such as endosomes, lysosomes, and secretory vesicles. Briefly, cells (2.4×10⁵) were seeded in six-well plates overnight and then treated with the ANE or vehicle for 48 h. Cells were then harvested and stained with fluorescent dyes from a LYSO-ID® Green detection kit and measured using the NucleoCounter® NC-3000^TM system (ChemoMetec).

Wei P.L., Hung C.S., Lu H.H., Batzorig U., Huang C.Y, & Chang Y.J. (2021). Areca nut extract (ANE) inhibits the progression of hepatocellular carcinoma cells via activation of ROS production and activation of autophagy. International Journal of Medical Sciences, 18(15), 3452-3462.

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Quantifying Acidic Vesicles in Live Cells

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Acidic vesicles were detected in live cells using the Lyso-ID Green detection kit (Enzo Lifesciences, Farmingdale, NY, USA). Cells were seeded in 96-well plates or 12 well plates (90% confluent) and incubated with 4 μl/ml Lyso-ID in assay buffer supplemented with 2% (v/v) FCS at 37 °C for 30 min. Cells were washed with assay buffer and fluorescence measured on a plate reader at excitation 488 nm, emission 520 nm. Following reading, the buffer was aspirated, cells lysed overnight in 0.25M NaOH and protein concentration measured. Cell fluorescence was expressed as fluorescent units/mg protein.
For live cell imaging, cells were seeded on glass bottom dishes (ibidi, Germany) and incubated with 1μl/ml Lyso-ID in HBSS for 15 min. Cells were then washed with HBSS and imaged in HBSS.

Magalhaes J., Gegg M.E., Migdalska-Richards A., Doherty M.K., Whitfield P.D, & Schapira A.H. (2016). Autophagic lysosome reformation dysfunction in glucocerebrosidase deficient cells: relevance to Parkinson disease. Human Molecular Genetics, 25(16), 3432-3445.

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Quantifying PG-induced Lysosome Formation

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PG-induced lysosome formation was measured using the LYSO-ID Green detection kit (ENZ-51034, Enzo). The dye accumulates in acidic compartments, such as endosomes, lysosomes, and secretory vesicles. Briefly, 2.4×10⁵ cells were seeded in six-well plates overnight. After treatment with 80 μg/ml PG or vehicle for 48 h, cells were then harvested and stained with fluorescent dyes using the LYSO-ID Green detection kit and measured using the NucleoCounter NC-3000 system (ChemoMetec).

Wei P.L., Huang C.Y, & Chang Y.J. (2019). Propyl gallate inhibits hepatocellular carcinoma cell growth through the induction of ROS and the activation of autophagy. PLoS ONE, 14(1), e0210513.

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