Lysosomal Intracellular Activity Assay Kit (ab234622)
Key features and details
- Assay type: Cell-based
- Detection method: Fluorescent
- Platform: Flow cytometer, Fluorescence microscope
- Sample type: Adherent cells, Suspension cells
Overview
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Product name
Lysosomal Intracellular Activity Assay Kit -
Detection method
Fluorescent -
Sample type
Adherent cells, Suspension cells -
Assay type
Cell-based -
Species reactivity
Reacts with: Mammals, Other species -
Product overview
Lysosomal Intracellular Activity Assay Kit (ab234622) provides 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. The kit includes Cytochalasin D, a cell-permeable inhibitor of the lysosomal membrane V-ATPase proton pump, that serves as an anti-lysosomal experimental control. This easy-to-use, non-radioactive kit allows imaging and accurate measurement of de-quenching substrate in cultured cells.
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Notes
This product is manufactured by BioVision, an Abcam company and was previously called K448 Lysosomal Intracellular Activity Assay Kit. K448-50 is the same size as the 50 test size of ab234622.
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.
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Platform
Flow cytometer, Fluorescence microscope
Properties
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Storage instructions
Store at -20°C. Please refer to protocols. -
Components 50 tests 100X Cytochalasin D 1 x 50µl Buffer I 1 x 1.8ml Self-Quenched Substrate 1 vial
Images
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Release of self-Quenched Substrate in U937 cells. 1x106 U937 cells were pretreated with or without 1X Cytochalasin D for 1 hr. After pretreatment, remove the medium, cells were incubated with Self-Quenced Substrate, and the same concentration of Cytochalasin D for another 1 hr in medium with 0.5% FBS according to kit’s protocol. (A) Comparison of histograms from flow analysis showing the inhibition of De-Quenching of Substrate by Cytochalasin D in U937 cells (Black: negative control cells; Pink: in the presence of 1X Cytochalasin D; Green: without 1X Cytochalasin D).
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1x106 U937 cells were pretreated with vehicle or 1X Cytochalasin D for 1 hour. 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. 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).
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1x106 U937 cells were pretreated with vehicle or 1X Cytochalasin D for 1 hour. 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. Lysosomal staining with Lysosomal Associated Membrane Protein 1 (Lamp-1 RFP, a lysosomal marker). Cells were stained with nuclear dye for 10 minutes, washed with 1X PBS and mounted on the slide. Images were taken using a fluorescence microscope with a 60X objective lens.
Datasheets and documents
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SDS download
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Datasheet download
References (9)
ab234622 has been referenced in 9 publications.
- Suda M et al. Glycoprotein nonmetastatic melanoma protein B regulates lysosomal integrity and lifespan of senescent cells. Sci Rep 12:6522 (2022). PubMed: 35444208
- Wang M et al. Domain fusion TLR2-4 enhances the autophagy-dependent clearance of Staphylococcus aureus in the genetic engineering goat. Elife 11:N/A (2022). PubMed: 35762728
- Wakao S et al. Phagocytosing differentiated cell-fragments is a novel mechanism for controlling somatic stem cell differentiation within a short time frame. Cell Mol Life Sci 79:542 (2022). PubMed: 36203068
- Busnelli M et al. Aortic Gene Expression Profiles Show How ApoA-I Levels Modulate Inflammation, Lysosomal Activity, and Sphingolipid Metabolism in Murine Atherosclerosis. Arterioscler Thromb Vasc Biol 41:651-667 (2021). PubMed: 33327742
- Ketterer S et al. Cathepsin D deficiency in mammary epithelium transiently stalls breast cancer by interference with mTORC1 signaling. Nat Commun 11:5133 (2020). PubMed: 33046706
- Ghosh S et al. ß-Coronaviruses Use Lysosomes for Egress Instead of the Biosynthetic Secretory Pathway. Cell 183:1520-1535.e14 (2020). PubMed: 33157038
- Ho PW et al. Age-dependent accumulation of oligomeric SNCA/a-synuclein from impaired degradation in mutant LRRK2 knockin mouse model of Parkinson disease: role for therapeutic activation of chaperone-mediated autophagy (CMA). Autophagy 16:347-370 (2020). PubMed: 30983487
- Rijal R et al. Polyphosphate is an extracellular signal that can facilitate bacterial survival in eukaryotic cells. Proc Natl Acad Sci U S A 117:31923-31934 (2020). PubMed: 33268492
- Pareja F et al. Loss-of-function mutations in ATP6AP1 and ATP6AP2 in granular cell tumors. Nat Commun 9:3533 (2018). PubMed: 30166553