DEPTOR/DEPDC6 Antibody - BSA Free

Novus Biologicals | Catalog # NBP1-49674

Novus Biologicals
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Key Product Details

Validated by

Knockout/Knockdown

Species Reactivity

Validated:

Human, Mouse, Rat, Bovine

Cited:

Human, Mouse, Rat, Bovine

Applications

Validated:

Knockout Validated, Immunohistochemistry, Immunohistochemistry-Paraffin, Western Blot, Immunocytochemistry/ Immunofluorescence

Cited:

Immunohistochemistry-Paraffin, Western Blot, Immunocytochemistry/ Immunofluorescence

Label

Unconjugated

Antibody Source

Polyclonal Rabbit IgG

Format

BSA Free
View all DEPTOR/DEPDC6 Primary Antibodies »

Product Specifications

Immunogen

A genomic peptide made to an internal region of the human DEPTOR protein (within residues 10-200). [Swiss-Prot: Q8TB45]

Reactivity Notes

Rat reactivity reported in scientific literature (PMID: 31176743).

Localization

Intracellular.

Clonality

Polyclonal

Host

Rabbit

Isotype

IgG

Scientific Data Images for DEPTOR/DEPDC6 Antibody - BSA Free

Western Blot: DEPTOR/DEPDC6 AntibodyBSA Free [NBP1-49674]

Western Blot: DEPTOR/DEPDC6 AntibodyBSA Free [NBP1-49674]

DEPTOR-DEPDC6-Antibody---BSA-Free-Western-Blot-NBP1-49674-img0012.jpg
Immunocytochemistry/ Immunofluorescence: DEPTOR/DEPDC6 Antibody - BSA Free [NBP1-49674]

Immunocytochemistry/ Immunofluorescence: DEPTOR/DEPDC6 Antibody - BSA Free [NBP1-49674]

Immunocytochemistry/Immunofluorescence: DEPTOR/DEPDC6 Antibody - BSA Free [NBP1-49674] - Antibody was tested in MCF7 cells with DyLight 488 (green). Nuclei and alpha-tubulin were counterstained with DAPI (blue) and DyLight 550 (red).
Immunohistochemistry: DEPTOR/DEPDC6 Antibody - BSA Free [NBP1-49674]

Immunohistochemistry: DEPTOR/DEPDC6 Antibody - BSA Free [NBP1-49674]

Immunohistochemistry: DEPTOR/DEPDC6 Antibody - BSA Free [NBP1-49674] - Staining of DEPTOR in mouse spleen using DAB with a hematoxylin counterstain.
Western Blot: DEPTOR/DEPDC6 AntibodyBSA Free [NBP1-49674]

Western Blot: DEPTOR/DEPDC6 AntibodyBSA Free [NBP1-49674]

Western Blot: DEPTOR/DEPDC6 Antibody - BSA Free [NBP1-49674] - Analysis of DEPDC6 in (A) human liver and (B) mouse liver.
Knockout Validated: DEPTOR/DEPDC6 Antibody - BSA Free [NBP1-49674]

Western Blot: DEPTOR/DEPDC6 Antibody - BSA Free [NBP1-49674]

DEPTOR-DEPDC6 Antibody - BSA Free-Knockout Validated-NBP1-49674-img0013.jpg

Applications for DEPTOR/DEPDC6 Antibody - BSA Free

Application
Recommended Usage

Immunocytochemistry/ Immunofluorescence

1:100

Immunohistochemistry

1:100

Immunohistochemistry-Paraffin

1:100

Western Blot

1:1000
Application Notes
In Western Blot, a band is seen at ~46 kDa. In ICC/IF, cytoplasmic staining was observed in serum starved HeLa cells. In IHC-P, staining was observed in the cytoplasm of mouse spleen. Prior to immunostaining paraffin tissues, antigen retrieval with sodium citrate buffer (pH 6.0) is recommended.

Reviewed Applications

Read 2 reviews rated 4 using NBP1-49674 in the following applications:

Formulation, Preparation, and Storage

Purification

Immunogen affinity purified

Formulation

PBS

Format

BSA Free

Preservative

0.02% Sodium Azide

Concentration

1.0 mg/ml

Shipping

The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below.

Stability & Storage

Store at 4C short term. Aliquot and store at -20C long term. Avoid freeze-thaw cycles.

Background: DEPTOR/DEPDC6

DEPTOR (DEP domain-containing mTOR-interacting protein), also called DEP domain-containing protein 6 (DEPDC6), is a part of mTORC1 (MTOR, MLST8, RPTOR, AKT1S1/PRAS40 and DEPTOR) and mTORC2 (MTOR, MLST8, PROTOR1, RICTOR, MAPKAP1 and DEPTOR) complexes. DEPTOR functions as a negative regulator of the mTORC1/mTORC2 signaling pathways, leading to inhibition of activity of both kinase complexes. DEPTOR phosphorylation can weaken the interaction with MTOR within mTORC1 and mTORC2 complexes. DEPTOR has been shown to act as a tumor suppressor via mTORC1/mTORC2 blocking to inhibit protein synthesis as well as AKT's proliferation and survival related signaling. However, in certain conditions, DEPTOR behaves as an oncogene by relieving the feedback inhibition from S6K1 to PI3K, thus causing AKT activation. The stability of DEPTOR is controlled by SCFbeta-TrCP E3 ubiquitin ligase, and S6K1 and RSK1 are kinases that phosphorylate DEPTOR for facilitating its binding to betaTrCP for subsequent degradation. DEPTOR is down-regulated in a number of malignancies, however, is overexpressed in a subset of multiple myelomas harbouring cyclin D1/D3 or c-MAF/MAFB translocations.

Long Name

DEP Domain Containing mTOR-interacting Protein

Alternate Names

DEP.6, DEPDC6

Entrez Gene IDs

64798 (Human); 97998 (Mouse)

Gene Symbol

DEPTOR

UniProt

Q8TB45 (Human), Q570Y9 (Mouse)

Additional DEPTOR/DEPDC6 Products

Product Documents for DEPTOR/DEPDC6 Antibody - BSA Free

Certificate of Analysis

To download a Certificate of Analysis, please enter a lot or batch number in the search box below.

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Product Specific Notices for DEPTOR/DEPDC6 Antibody - BSA Free

Manufactured by Genomic Antibody Technology™. GAT FAQs

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed for 1 year from date of receipt.

Citations for DEPTOR/DEPDC6 Antibody - BSA Free

Customer Reviews for DEPTOR/DEPDC6 Antibody - BSA Free (2)

4 out of 5
2 Customer Ratings
5 Stars
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4 Stars
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3 Stars
50%
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1 Stars
0%

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Customer Images

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  • DEPTOR/DEPDC6 Antibody
    Name: Anonymous
    Application: Western Blot
    Sample Tested: Adipose tissue
    Species: Mouse
    Verified Customer | Posted 10/23/2019
    Mouse white adipose tissue was homogenised and protein content was quantified by a BCA assay. Twenty micrograms of protein were resolved on a 4-12% Bis-Tris gel and transferred to nitrocellulose membranes. Membranes were probed with primary antibody DEPTOR diluted 1:1000 in 5% BSA, before incubation with Anti-rabbit secondary horseradish peroxidase-conjugated antibody 1:5000. Blots were visualised with Immobilon Western chemiluminescence HRP Substrate and imaged with Syngene chemiluminescence imaging system.
    DEPTOR/DEPDC6 Antibody - BSA Free NBP1-49674
  • Name: Anonymous
    Application: Western Blot
    Verified Customer | Posted 08/29/2014
    Full blot corresponding to control or DEPTOR knocked down in 3t3-L1 cells using lentiviruses expressing shRNA
    DEPTOR/DEPDC6 Antibody - BSA Free NBP1-49674

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Protocols

View specific protocols for DEPTOR/DEPDC6 Antibody - BSA Free (NBP1-49674):


Immunocytochemistry Protocol

Culture cells to appropriate density in 35 mm culture dishes or 6-well plates.

1. Remove culture medium and add 10% formalin to the dish. Fix at room temperature for 30 minutes.
2. Remove the formalin and add ice cold methanol. Incubate for 5-10 minutes.
3. Remove methanol and add washing solution (i.e. PBS). Be sure to not let the specimen dry out. Wash three times for 10 minutes.
4. To block nonspecific antibody binding incubate in 10% normal goat serum from 1 hour to overnight at room temperature.
5. Add primary antibody at appropriate dilution and incubate at room temperature from 2 hours to overnight at room temperature.
6. Remove primary antibody and replace with washing solution. Wash three times for 10 minutes.
7. Add secondary antibody at appropriate dilution. Incubate for 1 hour at room temperature.
8. Remove antibody and replace with wash solution, then wash for 10 minutes. Add Hoechst 33258 to wash solution at 1:25,0000 and incubate for 10 minutes. Wash a third time for 10 minutes.
9. Cells can be viewed directly after washing. The plates can also be stored in PBS containing Azide covered in Parafilm (TM). Cells can also be cover-slipped using Fluoromount, with appropriate sealing.

*The above information is only intended as a guide. The researcher should determine what protocol best meets their needs. Please follow safe laboratory procedures.


Immunohistochemistry-Paraffin Embedded Sections

Antigen Unmasking:
Bring slides to a boil in 10 mM sodium citrate buffer (pH 6.0) then maintain at a sub-boiling temperature for 10 minutes. Cool slides on bench-top for 30 minutes.

Staining:
1. Wash sections in deionized water three times for 5 minutes each.
2. Wash sections in wash buffer for 5 minutes.
3. Block each section with 100-400 ul blocking solution for 1 hour at room temperature.
4. Remove blocking solution and add 100-400 ul diluted primary antibody. Incubate overnight at 4C.
5. Remove antibody solution and wash sections in wash buffer three times for 5 minutes each.
6. Add 100-400 ul biotinylated diluted secondary antibody. Incubate 30 minutes at room temperature.
7. Remove secondary antibody solution and wash sections three times with wash buffer for 5 minutes each.
8. Add 100-400 ul Streptavidin-HRP reagent to each section and incubate for 30 minutes at room temperature.
9. Wash sections three times in wash buffer for 5 minutes each.
10. Add 100-400 ul DAB substrate to each section and monitor staining closely.
11. As soon as the sections develop, immerse slides in deionized water.
12. Counterstain sections in hematoxylin.
13. Wash sections in deionized water two times for 5 minutes each.
14. Dehydrate sections.
15. Mount coverslips.


Western Blot Protocol

1. Perform SDS-PAGE on samples to be analyzed, loading 40 ug of total protein per lane.
2. Transfer proteins to membrane according to the instructions provided by the manufacturer of the membrane and transfer apparatus.
3. Stain according to standard Ponceau S procedure (or similar product) to assess transfer success, and mark molecular weight standards where appropriate.
4. Rinse the blot.
5. Block the membrane using standard blocking buffer for at least 1 hour.
6. Wash the membrane in wash buffer three times for 10 minutes each.
7. Dilute the rabbit anti-Deptor primary antibody (NBP1-49674) in blocking buffer and incubate 1 hour at room temperature.
8. Wash the membrane in wash buffer three times for 10 minutes each.
9. Apply diluted rabbit-IgG HRP-conjugated secondary antibody in blocking buffer (as per manufacturers instructions) and incubate 1 hour at room temperature.
10. Wash the blot in wash buffer three times for 10 minutes each (this step can be repeated as required to reduce background).
11. Apply the detection reagent of choice in accordance with the manufacturers instructions.

*Note: Tween-20 can be added to the blocking or antibody dilution buffer at a final concentration of 0.05-0.2%.

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