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ProteoStat® Aggresome detection kit for flow cytometry and fluorescence microscopy

Robust, quantitative detection of aggresomes relevant to the study of neurodegenerative disease, liver disease and toxicology
 
ENZ-51035-0025 25 tests 100.00 USD
 
ENZ-51035-K100 100 tests 274.00 USD
 
Replaces Prod. #: ENZ-51038

  • Provides a sensitive cell-based assay of drug responsiveness to identify inhibitors relevant to neurodegenerative disease in an authentic cellular context
  • Reliable and simple assay doesn’t require non-physiological protein mutations or genetically engineered cell lines
  • Validated under a wide range of conditions and with small molecule modulators demonstrating suitability for screening compounds of potential therapeutic value
  • Fixed-cell assay is optimized for antibody co-localization studies
  • Easily quantifies aggresome and related inclusion bodies by flow cytometry
  • Useful for the study of neurodegenerative diseases; Liver disease; Toxicology studies and much more
Aggresomes are inclusion bodies that form when the ubiquitin–proteasome machinery is overwhelmed with aggregation-prone proteins. Typically, an aggresome forms in response to some cellular stress, such as hyperthermia, viral infection, or exposure to reactive oxygen species. Aggresomes may provide a cytoprotective function by sequestering the toxic, aggregated proteins and may also facilitate their ultimate elimination from cells by autophagy.
Certain cellular inclusion bodies associated with human disease are thought to arise from an aggresomal response including Alzheimer’s disease as it relates to intracellular and extracellular aggregates, Lewy bodies associated with neurons in Parkinson's disease, Mallory bodies associated with liver cells in alcoholic liver disease and Hyaline inclusion bodies associated with astrocytes in amyotrophic lateral sclerosis (ALS).
ProteoStat® Aggresome Detection kit provides a rapid, specific and quantitative approach to identify inhibitors relevant to neurodegenerative disease in an authentic cellular context. This fixed-cell assay doesn’t require non-physiological protein mutations or genetically engineered cell lines. ProteoStat® dye has been validated under a wide range of conditions and with small molecule modulators, demonstrating suitability for screening compounds of potential therapeutic value, and optimized for antibody co-localization studies to identify interactions between aggregated protein cargo and the various proteins implicated in autophagy and aggresome formation.
ENZ-51035 control
Aggresomes with HeLa cells, treated for 12 hours with 5µM MG-132 (right), detected by ProteoStat® Aggresome dye (red) and counterstained with Hoechst 33342. Control panel on left is untreated.
ENZ-51035 co-locolize
Aggresomes with HeLa cells, previously treated for 12 hours with 5µM MG-132, detected by ProteoStat® Aggresome dye (upper left, appearing here in green) showing co-localization with Alexa Fluor® 647-Tubulin antibody (lower left, appearing in red) and composite image (right), as observed by fluorescence microscopy.
ENZ-51035 02
Aggresomes within HeLa cells, previously treated for 12 hours with 5μM MG-132, detected by ProteoStat® aggresome dye (upper left) showing co-localization with fluorescein-p62 antibody (upper right) and composite image (lower center), as observed by fluorescence microscopy.
Please mouse over
ENZ-51035 control ENZ-51035 co-locolize ENZ-51035 02

Product Specification

Applications:FC, Fluorescence microscopy, Fluorescent detection
 
Application Notes:The ProteoStat®Aggresome detection kit assay is potentially applicable to the identification of small molecules that inhibit aggresome formation as well as immuno-localization studies between aggregated protein cargo and the various proteins implicated in aggresome formation, such as histone deacetylase 6, parkin, ataxin-3, dynein motor complex and ubiquilin-1.
 
Quality Control:A sample from each lot of ProteoStat® Aggresome detection kit is used to stain Jurkat cells and analyzed by flow cytometry, using the procedures described in the user manual. The aggresome activity factor (AAF) values for the samples are greater than 25. 
 
Quantity:For ENZ-51035-0025:
25 flow cytometry assays or 50 microscopy assays

For ENZ-51035-K100:
100 flow cytometry assays or 200 microscopy assays
 
Use/Stability:With proper storage, the kit components are stable for one year from date of receipt.
 
Handling:Protect from light. Avoid freeze/thaw cycles.
 
Short Term Storage:-20°C
 
Long Term Storage:-80°C
 
Kit/Set Contains:ProteoStat®Aggresome detection reagent 
Hoechst 33342 Nuclear stain
Proteasome Inhibitor (MG-132)
10X Assay Buffer
 
Background / Technical Information:Enzo and ProteoStat® are trademarks of Enzo Life Sciences, Inc. Several of Enzo’s products and product applications are covered by US and foreign patents and patents pending.

Application Note: Cell-Based Screening of Focused Bioactive Compound Libraries: Assessing Small Molecule Modulators of the Canonical Wnt Signaling and Autophagy-Lysosome Pathways.
 

Product Literature References

Direct visualization of HIV-enhancing endogenous amyloid fibrils in human semen: S. M. Usmani, et al.; Nat. Commun. 5, 3508 (2014), Application: Amyloid detection in semen, Abstract;
Distinct patterns of HSP30 and HSP70 degradation in Xenopus laevis A6 cells recovering from thermal stress: S. Khan, et al.; Comp. Biochem. Physiol. A Mol. Integr. Physiol. 168, 1 (2014), Application(s): Detection of aggresomes in Xenopus laevis cells using fluorescence microscopy, Abstract;
Novel estradiol analogue induces apoptosis and autophagy in esophageal carcinoma cells: E. Wolmarans, et al.; Cell Mol. Biol. Lett. 19, 98 (2014), Abstract;
Preconditioning stimulus of proteasome inhibitor enhances aggresome formation and autophagy in differentiated SH-SY5Y cells: Y. Bang, et al.; Neurosci. Lett. 566, 263 (2014), Abstract;
Protein expression pattern of PAWP in bull spermatozoa is associated with sperm quality and fertility following artificial insemination: C.E. Kennedy, et al.; Mol. Reprod. Dev. 81, 436 (2014), Abstract;
Aldosterone and angiotensin II induce protein aggregation in renal proximal tubules: M.U. Cheema, et al.; Physiol. Rep. 1, e00064 (2013), Application(s): Labeling of kidney homogenates, labeled particles sorted by flow cytometry and identification by LC-MS/MS , Abstract; Full Text
Covalent and allosteric inhibitors of the ATPase VCP/p97 induce cancer cell death: P. Magnaghi, et al.; Nat. Chem. Biol. 9, 548 (2013), Application(s): Detection of aggresomes in human colon carcinoma HCT116 cells using fluorescence microscopy, Abstract;
Environmental stresses induce misfolded protein aggregation in plant cells in a microtubule-dependent manner: Y. Nakajima, et al.; Int. J. Mol. Sci. 14, 7771 (2013), Application(s): Detection of aggresomes using fluorescence microscopy, Abstract; Full Text
In vitro changes in mitochondrial potential, aggresome formation and caspase activity by a novel 17-β-estradiol analogue in breast adenocarcinoma cells: D.S. Nkandeu, et al.; Cell. Biochem. Funct. (2013), Abstract;
Increased generation of cyclopentenone prostaglandins after brain ischemia and their role in aggregation of ubiquitinated proteins in neurons: H. Liu, et al.; Neurotox. Res. 24, 191 (2013), Abstract;
Macrolide antibiotics block autophagy flux and sensitize to bortezomib via endoplasmic reticulum stress-mediated CHOP induction in myeloma cells: S. Moriya, et al.; Int. J. Oncol. 42, 1541 (2013), Application(s): Detection of aggresomes using flow cytometry, Abstract; Full Text
Mst1 inhibits autophagy by promoting the interaction between Beclin1 and Bcl-2: Y. Maejima, et al.; Nat. Med. 19, 1478 (2013), Application(s): Detection of aggresomes in mouse heart sections using fluorescence microscopy, Abstract;
N-terminally truncated forms of human cathepsin F accumulate in aggresome-like inclusions: B. Jeric, et al.; Biochim. Biophys. Acta 1833, 2254 (2013), Application(s): Detection of aggresomes using fluorescence microscopy, Abstract;
The ubiquitin proteasome system regulates the stability and activity of the glucose sensor glucokinase in pancreatic beta cells: A. Hofmeister-Brix, et al.; Biochem. J. (2013), Abstract; Full Text
VCP Phosphorylation-Dependent Interaction Partners Prevent Apoptosis in Helicobacter pylori-Infected Gastric Epithelial Cells: C.C. Yu, et al.; PLoS One 8, e55724 (2013), Application(s): Aggresome detection in AGS human gastric epithelial cells, Abstract; Full Text
Zerumbone, an electrophilic sesquiterpene, induces cellular proteo-stress leading to activation of ubiquitin-proteasome system and autophagy: K. Ohnishi, et al.; BBRC 430, 616 (2013), Application(s): Aggresome detection in mouse hepatocytes, Abstract;
Autophagy in idiopathic pulmonary fibrosis: A.S. Patel, et al.; PLoS One 7, e41394 (2012), Application(s): Detection of aggresomes in lung tissue sections using fluorescence microscopy, Abstract; Full Text
Decreased proteasomal activity causes age-related phenotypes and promotes the development of metabolic abnormalities: U. Tomaru, et al.; Am. J. Pathol. 180, 963 (2012), Abstract;
Mutations in the area composita protein αT-catenin are associated with arrhythmogenic right ventricular cardiomyopathy: J. van Hengel, et al.; Eur. Heart J. 34, 201 (2012), Abstract;
Quantitative analysis of α-synuclein solubility in living cells using split GFP complementation: A. Kothawala, et al.; PLoS One 7, e43505 (2012), Application(s): Aggresome detection in HeLa cells, Abstract; Full Text
Multiple aggregates and aggresomes of C-terminal truncated human αA-crystallins in mammalian cells and protection by αB-crystallin: I. Raju, et al.; PLoS One 6, e19876 (2011), Application(s): Aggresome detection in HeLa cells, Abstract; Full Text
Novel Cell- and Tissue-Based Assays for Detecting Misfolded and Aggregated Protein Accumulation Within Aggresomes and Inclusion Bodies: D. Shen, et al.; Cell Biochem. Biophys. 60, 173 (2011), Abstract;

General Literature References

Inhibitors of protein aggregation and toxicity: H. Amijee, et al.; Biochem. Soc. Trans. 37, 692 (2009), Full Text
Autophagy-mediated clearance of aggresomes is not a universal phenomenon: E. Wong, et al.; Hum. Mol. Genet. 17, 2570 (2008), Full Text
Proteostasis regulators and pharmacologic chaperones synergise to correct protein misfolding diseases: T-W Mu, et al.; Cell 134, 769 (2008), Full Text
Inhibitors of the proteasome suppress homologous DNA recombination in mammalian cells: Y. Murakawa, et al.; Cancer Res. 67, 8536 (2007), Full Text
p62SQSTM1 forms protein aggregates degraded by autophagy and has a protective effect on huntingtin-induced cell death: G. Bjørkøy, et al.; J.Cell Biol. 171, 603 (2005), Full Text
Therapeutic effects of cystamine in a murine model of Huntington's disease: A. Dedeoglu, et al.; J. Neurosci. 22, 8942 (2002), Full Text

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