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Autophagy Antibody Sampler Kit

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RUO
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Please refer to individual product datasheets.
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Autophagy_Antibody_Sampler_Kit_1_101918
Western blot of anti-Ubiquitin antibody (clone P4G7). Lane 1: Molecular weight marker; Lane 2: Poly-His tagged human ubiquitin protein. The blot was incubated with the primary antibody at 10 µg/ml for 24 hours at 4°C, followed by incubation with HRP labeled goat anti-mouse secondary antibody and visualization with a chemiluminescence detection system.
  • Autophagy_Antibody_Sampler_Kit_1_101918
    Western blot of anti-Ubiquitin antibody (clone P4G7). Lane 1: Molecular weight marker; Lane 2: Poly-His tagged human ubiquitin protein. The blot was incubated with the primary antibody at 10 µg/ml for 24 hours at 4°C, followed by incubation with HRP labeled goat anti-mouse secondary antibody and visualization with a chemiluminescence detection system.
  • Autophagy_Antibody_Sampler_Kit_2_101918
    Western blot of anti-Rab7A antibody (clone W16034A). The blot was incubated with 2µg/ml of the primary antibody overnight at 4°C, followed by the incubation with horseradish peroxidase labeled goat anti-mouse secondary antibody. Enhanced chemiluminescence was used as the detection system. M: Molecular weight marker; brain lysates: 20 µg; recombinant proteins: 10 ng.
  • Autophagy_Antibody_Sampler_Kit_3_101918
    Western blot of anti-Beclin-1 antibody (clone O93F3) and isotype-matched IgG1 control. Blots were incubated with 5µg/ml (left) or 0.5µg/ml (middle) anti-Beclin-1 antibody or IgG1 isotype control antibody (right) overnight at 4°C, followed by the incubation with horseradish peroxidase labeled goat anti-mouse secondary antibody. Enhanced chemiluminescence was used as the detection system. M: Molecular weight marker; brain lysates: 20 µg. The anti-Histone-H3 antibody was used as a loading control.
  • Autophagy_Antibody_Sampler_Kit_4_101918
    Western blot of anti-ATG5 antibody (clone 177.19). The blot was incubated with the primary antibody at 0.2 µg/ml overnight at 4°C, followed by the incubation with horseradish peroxidase labeled goat anti-mouse secondary antibody (Cat. No. 405306). Enhanced chemiluminescence was used as the detection system. Lysates: 10 µg. Recombinant human ATG5: 5 ng.
  • Autophagy_Antibody_Sampler_Kit_5_101918
    ICC staining of anti-ATG5 antibody (clone 177.19) on Hela cells. The cells were fixed with 4% PFA, permeabilized with 0.1% Triton X-100, and blocked with 2% normal goat serum and 0.02%BSA. The cells were then stained with 0.5 µg/ml of the primary antibody overnight at 4°C, followed by incubation with Alexa Fluor® 594 goat anti-Mouse IgG (Cat. No. 405326) for one hour at room temperature. Cells were counterstained with Phalloidin™ Green 488 (Cat. No. 424201) and DAPI to visualize actin filaments and nuclei, respectively. The images were captured with a 60X objective. Scale bar: 50 µm.
  • Autophagy_Antibody_Sampler_Kit_6_101918
    SH-SY5Y neuroblastoma cells were fixed with 4% paraformaldehyde, and then permeabilized with 0.1% Triton x-100 for 20 minutes. Cells were blocked with 2% normal goat serum for 30 minutes at room temperature, and then incubated with anti-LC3 antibody (clone A15143K) at 1 µg/mL for three hours at room temperature. Alexa Fluor® 488 conjugated anti-mouse IgG (green) was used as secondary antibody at 1 µg/mL. Cells were then incubated with anti-Tubulin β3 (TUBB3) antibody (clone TUJ1) Alexa Fluor® 647 (pink) at 1 µg/mL overnight at 4°C. Nuclei were stained with Hoechst 33342 (blue) at 5 µg/mL for 10 minutes at room temperature. Images were captured with a 60X objective.
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Description

Autophagy is a regulated process for protein degradation and turnover of damaged organelles to maintain cellular homeostasis. Under conditions such as stress or starvation, autophagy contributes to the breakdown of proteins and cellular compartments to generate amino acids required for the synthesis of proteins essential for cell survival. Dysregulation of autophagy or mutations in proteins involved in this pathway have been shown to contribute to neurodegenerative disorders such as Parkinson’s disease. The Autophagy Antibody Sampler Kit provides flexibility for sampling and detection of key targets in the autophagy pathway.

Product Details
Technical data sheet

Kit Contents

Kit Contents
Specificity Format Clone Size Reactivity Isotype
Anti-Ubiquitin Purified P4G7 25 μg Human, Extensive (Yeast to Human) Mouse IgG1, κ
Anti-Rab7A Purified W16034A 25 μg Human, Rat, Mouse Rat IgG2a, κ
Anti-Beclin-1 Purified O93F3 25 μg Human, Rat, Mouse Mouse IgG1
Anti-ATG5 Purified 177.19 25 μg Human, Mouse, Rat Mouse IgG1, κ
 Anti-LC3 Purified A15143K 25 μg Human, Mouse Mouse IgG2b, κ

* For detailed information about each specificity, please refer to the datasheets of the individual products. 

Product Details

Formulation
Please refer to individual product datasheets of the purified formats for details.
Preparation
All antibodies in this kit were purified by affinity chromatography.
Storage & Handling
Upon receipt, store undiluted at 2-8°C.
Application

WB, IHC-P - Quality tested
ICC - Verified
Recommended Usage

Each lot of antibodies in this kit is quality control tested by Western blotting. For Western blotting, the suggested uses of these reagents are as follows:

Anti-Ubiquitin: 1.0 - 10 µg/ml
Anti-Rab7A: 0.001 - 0.005 µg/ml
Anti-Beclin-1: 0.5 - 5.0 µg/ml
Anti-ATG5: 0.1 - 1.0 µg/ml

For immunohistochemistry, the suggested uses of these reagents are as follows:

Anti-ATG5: 0.2 - 1.0 µg/ml
Anti-LC3: 0.5 - 1.0 µg/ml

It is recommended that the reagent be titrated for optimal performance for each application.

Application Notes

For verified or reported applications for these antibodies, please see individual product datasheets.

Antigen Details

Biology Area
Cell Biology, Neurodegeneration, Neuroscience, Protein Trafficking and Clearance
Antigen References
  1. Uchiyama Y, et al. 2008. Histochem. Cell Biol. 129:407.
  2.  Wu H, et al. 2014. Int. J. Biol. Sci. 10:1072.
  3. Subramani S, Malhotra V. 2013. EMBO Rep. 14(2):143.
  4. Cao Y, et al. 2007. Cell. Research 17:839.
  5. Lee J, et al. 2012. Biochem. J. 441(2):523.
  6. Martini-Stoica H, et al. 2016. Trends Neurosci. 39(4)221.
  7. Guerra F, Bucci C. 2016. Cells. 5(3).
  8. Hatanaka H, et al. 2009. J. Biol. Chem. 23:15448.
Gene ID
9474 View all products for this Gene ID 8678 View all products for this Gene ID 81631 View all products for this Gene ID 7879 View all products for this Gene ID 7316 View all products for this Gene ID

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Go To Top Version: 1    Revision Date: 10/22/2018

For Research Use Only. Not for diagnostic or therapeutic use.

 

This product is supplied subject to the terms and conditions, including the limited license, located at www.biolegend.com/terms) ("Terms") and may be used only as provided in the Terms. Without limiting the foregoing, BioLegend products may not be used for any Commercial Purpose as defined in the Terms, resold in any form, used in manufacturing, or reverse engineered, sequenced, or otherwise studied or used to learn its design or composition without express written approval of BioLegend. Regardless of the information given in this document, user is solely responsible for determining any license requirements necessary for user’s intended use and assumes all risk and liability arising from use of the product. BioLegend is not responsible for patent infringement or any other risks or liabilities whatsoever resulting from the use of its products.

 

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