14-3-3 gamma [ac Val2] Antibody (KC21) - BSA Free

Novus Biologicals | Catalog # NB100-406

Novus Biologicals
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Key Product Details

Validated by

Biological Validation

Species Reactivity

Human

Applications

Western Blot, Immunocytochemistry/ Immunofluorescence, Simple Western

Label

Unconjugated

Antibody Source

Monoclonal Mouse IgG2A Clone # KC21

Format

BSA Free
View all 14-3-3 gamma Primary Antibodies »

Product Specifications

Immunogen

N-terminal fragment of human 14-3-3 gamma where the N-terminal Val was acetylated [UniProt# P61981]

Modification

ac Val2

Localization

Cytoplasmic

Specificity

This antibody is specific for the naturally occurring human form of 14-3-3 gamma, where the N-terminal Met is removed, resulting in an acetylated N-terminal Val. Unprocessed (non-modified) 14-3-3 gamma is not recognized by this antibody.

Clonality

Monoclonal

Host

Mouse

Isotype

IgG2A

Theoretical MW

33 kDa.
Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.

Scientific Data Images for 14-3-3 gamma [ac Val2] Antibody (KC21) - BSA Free

Western Blot: 14-3-3 gamma [ac Val2] Antibody (KC21) [NB100-406] - (1) HeLa, (2) PC12 and (3) NIH-3T3 whole cell lysates were separated by SDS-PAGE on a 12% gel and transferred to PVDF membrane. The membrane was probed with Anti-14-3-3 gamma [Ac Val2] antibody at 2 ug/ml for 1 hour, followed by incubation a 1:10,000 dilution of anti-rabbit HRP. The protein was detected at approximately 28 kDa after incubation with SuperSignal West Pico Chemiluminescent substrate.
Immunocytochemistry/Immunofluorescence: 14-3-3 gamma [ac Val2] Antibody (KC21) [NB100-406] - 14-3-3 gamma [ac Val2] antibody was tested in SH-SY5Y cells with DyLight 488 (green). Nuclei and alpha-tubulin were counterstained with DAPI (blue) and DyLight 550 (red).
Western Blot: 14-3-3 gamma [ac Val2] Antibody (KC21) [NB100-406] - 14-3-3 gamma detected in lysates using NB100-406. Lane 1: HeLa cell lysates, Lanes 2 and 3: bengamide treated lysates (8h and 24h, respectively).
Simple Western: 14-3-3 gamma [ac Val2] Antibody (KC21) [NB100-406] - Lane view shows a specific band for 14-3-3 Gamma in 1.0 mg/ml of HeLa lysate. This experiment was performed under reducing conditions using the 12-230kDa separation system.
14-3-3 gamma [ac Val2] Antibody (KC21) - BSA Free Immunocytochemistry/Immunofluorescence: 14-3-3 gamma [ac Val2] Antibody (KC21)

Immunocytochemistry/Immunofluorescence: 14-3-3 gamma [ac Val2] Antibody (KC21)

14-3-3 gamma was detected in immersion fixed HeLa human cervix adenocarcinoma cell line using Mouse anti-14-3-3 gamma Protein G Purified Monoclonal Antibody (Catalog # NB100-406) at 1.0 µg/mL overnight at 4C.  Cells were stained using DyLight 488-conjugated Anti-Mouse IgG (H+L) Cross-Absorbed Secondary Antibody (green), and counterstained with DAPI (blue). Cells were imaged using a 100X objective and digitally deconvolved.

Applications for 14-3-3 gamma [ac Val2] Antibody (KC21) - BSA Free

Application
Recommended Usage

Immunocytochemistry/ Immunofluorescence

1:500

Simple Western

1:12.5

Western Blot

1:1000-1:3000

Formulation, Preparation, and Storage

Purification

Protein G 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: 14-3-3 gamma

14-3-3 proteins are highly conserved and ubiquitously expressed proteins present as multigene families in most of the living organisms wherein they play a key role of bringing signal transduction to completion, and implicates in regulating several cellular processes including apoptosis, cell-cycle checkpoints and nutrient-sensing signaling pathways. 14-3-3 proteins have several isoforms which include -beta, -gamma, -epsilon, -sigma, -zeta, -tau, and -eta. 14-3-3 proteins can form homo- or heterodimers which interacts with a wide variety of proteins such as transcription factors, enzymes/kinases, cytoskeletal proteins, phosphatases, and other signaling molecules. 14-3-3 interactions with its targets occurs primarily via phospho-Ser/Thr motif, however, binding to divergent phospho-Ser/Thr motifs, as well as phosphorylation-independent interactions have been shown. 14-3-3 binding masks specific sequences of target proteins, thereby altering their localization, phosphorylation state, stability, and molecular interactions, and also induces target protein conformational changes which results in altered target protein function. Distinct temporal and spatial expression patterns of 14-3-3 isoforms have been observed in developmental and disease signaling which underlies their different functions despite sequence similarities.

Alternate Names

1433 gamma, YWHAG

Entrez Gene IDs

7532 (Human)

Gene Symbol

YWHAG

UniProt

P61981

Additional 14-3-3 gamma Products

Product Documents for 14-3-3 gamma [ac Val2] Antibody (KC21) - BSA Free

Certificate of Analysis

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Product Specific Notices for 14-3-3 gamma [ac Val2] Antibody (KC21) - BSA Free

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.

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Protocols

View specific protocols for 14-3-3 gamma [ac Val2] Antibody (KC21) - BSA Free (NB100-406):

14-3-3 gamma [ac Val2] Antibody (KC21):
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.

14-3-3 gamma [ac Val2] Antibody (KC21):
1. Load protein on gel (ie: ~30 ug of HeLa whole cell control lysate) and run.
2. Transfer protein to nitrocellulose (Schleicher&Schuell, cat# BA83).
3. Block the membrane with 5% milk for 30 minutes at room temperature.
4. Incubate the membrane with anti-14-3-3 gamma [cat# NB 100-406] (1:1,000), overnight at 4C or 2 h at room temperature. Milk or PBS-T are good for this dilution.
5. Wash the membrane 3 times, 10 minutes per wash in PBS with 0.1% Tween-20 (PBS-T).
6. Incubate with secondary antibody for 30 minutes at room temperature. Use milk in PBS-T as a diluent.
7. Rinse the membrane 2 times with deionized water and place in an ECL working solution.
8. Expose to the film.

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