Lysosomal Intracellular Activity Assay Kit (BN00690)

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BN00690
  • Lysosomal Intracellular Activity Assay Kit (BN00690)
  • Lysosomal Intracellular Activity Assay Kit (BN00690)
  • Lysosomal Intracellular Activity Assay Kit (BN00690)
€625

Description

ELISA Kit Technical ManualMSDS

Lysosomal Intracellular Activity Assay Kit

Lysosomes are membrane-bound organelles important for various cellular processes. They contain hydrolytic enzymes that are utilized in the metabolism of some biomolecules. The extracellular cargo (e.g. nutrients toxins) binds to the cell membrane and is subsequently transported into membrane-bound endosomes for further degradation by lysosomes while intracellular components are transported to lysosomes through autophagy. Lysosomal dysfunction is associated with many human conditions such as aging and neurodegenerative disease. Although the intracellular activity of lysosome is difficult to measure in living cells, Assay Genie has developed a proprietary Lysosome-Specific Self-Quenched Substrate which has low background fluorescence, high signal to background ratio and is pH insensitive. The substrate, acting as endocytic cargo, can be taken up by cells and degraded within an endo-lysosomal vesicle. The fluorescent signal is recovered from the Self-Quenched Substrate. The fluorescence signal, generated by degradation, is proportional to the intracellular lysosomal activity and can be measured using a fluorescence microscopy and/or flow cytometry. Lysosomal Intracellular Activity Assay Kit (Cell-Based) includes Cytochalasin D, a cell-permeable inhibitor of endocytosis that serves as an experimental control. This easy-to-use, non-radioactive kit allows imaging and accurate measurement of de-quenching substrate in cultured cells

Figure: Release of self-Quenched Substrate in U937 cells. 1x10 6 U937 cells were pretreated with vehicle or 1X Cytochalasin D for 1 hr. After pre-treatment, cells were incubated with Self-Quenched Substrate and the same concentration of Cytochalasin D for additional hour in medium supplemented with 0.5% FBS according to kit’s protocol. Panel A: Comparison of histograms from flow analysis showing the inhibition of De-Quenching of Substrate by Cytochalasin D. Unstained cells (black); experimental control (green) in the presence of 1X Cytochalasin D; Positive control (pink) without 1X Cytochalasin D. Panel B: Images of U937 cells obtained using fluorescence microscope. Top: positive control cells treated Self-quenched substrate only. Bottom: negative control cells treated with 1X Cytochalasin D. U937 cells showing the release of Self-quenched substrate in the endocytotic vesicle (punctured pattern). Panel C: Lysosomal staining with Lysosomal Associated Membrane Protein 1 (Lamp-1 RFP, a lysosomal marker). Cells were stained with nuclear dye for 10 min, washed with 1X PBS and mounted on the slide. Images were taken using a fluorescence microscope with a 60X objective lens.

Key Information Description

Product SKU

BN00690

Size

50 assays

Kit Summary

  • Detection method- Flow cytometry and Fluorescence Microscopy
  • Species reactivity- Mammalian
  • Applications -Measurement of Intracellular lysosomal activity and Screening compounds with anti-lysosomal activity

Detection Method

Flow Cytometry (FL1)/ Microscopy (Ex: 488 nm)

Species Reactivity

Mammalian

Applications

  • Measurement of lysosomal intracellular activity.
  • Elucidation of the mechanisms of endocytic pathway in living cells.
  • Screening compounds with anti-lysosomal intracellular activity

Features and Benefits

Measurement of lysosomal intracellular activity.

Kit Component

  • Polyamine Assay Buffer
  • Polyamine Probe (in DMSO)
  • Polyamine Enzyme Mix
  • Polyamine Developer
  • Sample Clean-Up Mix
  • Polyamine Standard

Storage Conditions

-20°C

Shipping Conditions

Gel Pack

USAGE

For Research Use Only! Not For Use in Humans.

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