GLI-3 Antibody

R&D Systems | Catalog # AF3690

R&D Systems

Key Product Details

Validated by

Biological Validation

Species Reactivity

Validated:

Human, Mouse

Cited:

Human, Mouse, Chicken, Drosophila, Nematode - Caenorhabditis elegans, Transgenic Mouse

Applications

Validated:

Western Blot, Immunocytochemistry, Chromatin Immunoprecipitation (ChIP)

Cited:

Immunohistochemistry, Immunohistochemistry-Paraffin, Western Blot, Immunocytochemistry, Immunoprecipitation, Chromatin Immunoprecipitation (ChIP), Bioassay, CUT&Tag

Label

Unconjugated

Antibody Source

Polyclonal Goat IgG
View all GLI-3 Primary Antibodies »

Product Specifications

Immunogen

E. coli-derived recombinant human GLI-3
Met1-Glu479
Accession # P10071

Specificity

Detects human and mouse GLI-3 in direct ELISAs and Western blots. In direct ELISAs and Western blots, less than 1% cross-reactivity with recombinant human (rh) GLI-1 and rhGLI-2 is observed.

Clonality

Polyclonal

Host

Goat

Isotype

IgG

Scientific Data Images for GLI-3 Antibody

Detection of Mouse GLI‑3 antibody by Western Blot.

Detection of Mouse GLI‑3 by Western Blot.

Western blot shows lysates of mouse embryo tissue. PVDF membrane was probed with 1 µg/mL of Goat Anti-Human/Mouse GLI-3 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF3690) followed by HRP-conjugated Anti-Goat IgG Secondary Antibody (Catalog # HAF019). A specific band was detected for GLI-3 at approximately 85 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 8.
Detection of GLI-3-regulated Genes antibody by Chromatin Immunoprecipitation.

Detection of GLI‑3-regulated Genes by Chromatin Immunoprecipitation.

Jurkat human acute T cell leukemia cell line treated with 50 ng/mL PMA and 200 ng/mL calcium ionomycin for 30 minutes was fixed using formaldehyde, resuspended in lysis buffer, and sonicated to shear chromatin. GLI-3/DNA complexes were immunoprecipitated using 5 µg Goat Anti-Human/Mouse GLI-3 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF3690) or control antibody (Catalog # AB-108-C) for 15 minutes in an ultrasonic bath, followed by Biotinylated Anti-Goat IgG Secondary Antibody (Catalog # BAF109). Immunocomplexes were captured using 50 µL of MagCellect Streptavidin Ferrofluid (Catalog # MAG999) and DNA was purified using chelating resin solution. Thegli-1promoter was detected by standard PCR.
GLI-3 antibody in HeLa Human Cell Line by Immunocytochemistry (ICC).

GLI‑3 in HeLa Human Cell Line.

GLI-3 was detected in immersion fixed HeLa human cervical epithelial carcinoma cell line using Human/Mouse GLI-3 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF3690) at 10 µg/mL for 3 hours at room temperature. Cells were stained using the NorthernLights™ 557-conjugated Anti-Goat IgG Secondary Antibody (yellow; Catalog # NL001) and counterstained with DAPI (blue). View our protocol for Fluorescent ICC Staining of Cells on Coverslips.
Detection of Mouse GLI-3 by Western Blot

Detection of Mouse GLI-3 by Western Blot

Conditional deletion of GATA6 does not significantly alter Gli3 processing in the hindlimb bud.(A) Western blot analysis of Gli3FL and Gli3R steady state levels in anterior (lanes 1 and 3) or posterior (lanes 2 and 4) halves of E11.5 hindlimb buds isolated from either control (lanes 1 and 2) or Prx1-Cre; GATA6fl/fl (lanes 3 and 4) mouse embryos. alpha –Tubulin is used as a loading control. (B) Quantification of Gli3FL/Gli3R ratios (by densitometry of Western Blots) in anterior or posterior halves of E11.5 hindlimb buds isolated from either littermate controls (lanes 1 and 2) or Prx1-Cre; GATA6fl/fl (lanes 3 and 4) animals, 12 embryos of each genotype were analyzed. Error bar indicates standard deviation, n = 12. Image collected and cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/24415953), licensed under a CC-BY license. Not internally tested by R&D Systems.
Detection of Mouse GLI-3 by Western Blot

Detection of Mouse GLI-3 by Western Blot

SufuT396I does not stabilize Gli3FL protein and reduces the processing of Gli3FL.(A) Western blotting of lysates prepared from SufuR146X/R146X at E9.5 and SufuT396I/T396I, SufuT396I/+, and wild-type embryos at E10.5 with anti-Gli3, anti-Sufu, and anti-actin antibodies. Each image presented in the Fig. is a representative of independent triplicated experiments. The full gel images are shown in S6A Fig. (B, C) Relative expression of Gli3FL (B) and Gli3REP (B and C). Western blotting was performed two times using lysates prepared from five wild-type and five SufuT396I/T396I embryos at E10.5 (S2A Fig.). Expression levels were quantified from the band intensity shown in S2A Fig. as relative values of the Gli3FL/actin and Gli3REP/actin expression ratios (B) and the direct ratio of Gli3REP/Gli3FL (C). (**) p < 0.01, two-tailed Student’s t-test. Error bars indicate the standard deviations. (D) Western blotting of cell lysates from wild-type and SufuT396I/T396I MEFs with indicated antibodies. The SufuT396I/T396I MEFs were electroporated with 10.0, 1.0, or 0.1 μg of the HA–Sufu construct (lane 3, 4, and 5, respectively), or 10 μg of the HA–SufuT396I construct (lane 6). The mobilities on SDS-PAGE of the wild-type Sufu (lane 3–5) and SufuT396I (lane 6) are identical. The complete gel images are shown in S6B Fig. This image is representative of two independent experiments. (E) Western blotting of cell lysates from Sufu−/− cells with indicated antibodies. The Sufu−/− cells were electroporated with a mixture of the Flag–Gli3 construct (6 μg) and the HA–Sufu construct (4.00, 1.33, 0.44, or 0.15 μg) for the wild-type Sufu cotransfection (lane 2 to 5, respectively), or a mixture of the Flag–Gli3 construct (6 μg) and the HA–SufuT396I construct (4 μg) for mutant Sufu cotransfection (lane 6). The complete gel images are shown in S6C Fig. This image is representative of two independent experiments. (F) Western blotting of immunoprecipitates or lysates from 293T cells transfected with exp
Detection of Mouse GLI-3 by Immunocytochemistry/ Immunofluorescence

Detection of Mouse GLI-3 by Immunocytochemistry/ Immunofluorescence

Analysis of Ptch receptor-dependent inhibition of Hh signaling. (A,C,E) Immunocytochemistry detection of Gli3 (A), Gli1 (C) and Gli2 (E) in the wild-type, Ptch1−/−, and Ptch2−/− MEFs. Scale bar 50 μm. (B,D,F) Quantification of the intensity of the nuclear GLI3 (B), GLI1 (D), and GLI2 (F) in wild-type, Ptch1−/−, and Ptch2−/− MEFs. **p ≤ 0.005, ***p ≤ 0.0005, ****p < 0.0001. (G) Western blot showing nuclear (N) and cellular (C) expression of Gli2 (FL = full length and RF = repressor form) in wild-type, Ptch1−/− and Ptch2−/− MEFs. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/35574464), licensed under a CC-BY license. Not internally tested by R&D Systems.
Detection of Mouse GLI-3 by Immunocytochemistry/ Immunofluorescence

Detection of Mouse GLI-3 by Immunocytochemistry/ Immunofluorescence

Analysis of Ptch receptor-dependent inhibition of Hh signaling. (A,C,E) Immunocytochemistry detection of Gli3 (A), Gli1 (C) and Gli2 (E) in the wild-type, Ptch1−/−, and Ptch2−/− MEFs. Scale bar 50 μm. (B,D,F) Quantification of the intensity of the nuclear GLI3 (B), GLI1 (D), and GLI2 (F) in wild-type, Ptch1−/−, and Ptch2−/− MEFs. **p ≤ 0.005, ***p ≤ 0.0005, ****p < 0.0001. (G) Western blot showing nuclear (N) and cellular (C) expression of Gli2 (FL = full length and RF = repressor form) in wild-type, Ptch1−/− and Ptch2−/− MEFs. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/35574464), licensed under a CC-BY license. Not internally tested by R&D Systems.
Detection of Mouse GLI-3 by Immunocytochemistry/ Immunofluorescence

Detection of Mouse GLI-3 by Immunocytochemistry/ Immunofluorescence

Analysis of Ptch receptor-dependent inhibition of Hh signaling. (A,C,E) Immunocytochemistry detection of Gli3 (A), Gli1 (C) and Gli2 (E) in the wild-type, Ptch1−/−, and Ptch2−/− MEFs. Scale bar 50 μm. (B,D,F) Quantification of the intensity of the nuclear GLI3 (B), GLI1 (D), and GLI2 (F) in wild-type, Ptch1−/−, and Ptch2−/− MEFs. **p ≤ 0.005, ***p ≤ 0.0005, ****p < 0.0001. (G) Western blot showing nuclear (N) and cellular (C) expression of Gli2 (FL = full length and RF = repressor form) in wild-type, Ptch1−/− and Ptch2−/− MEFs. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/35574464), licensed under a CC-BY license. Not internally tested by R&D Systems.
Detection of Mouse GLI-3 by Immunocytochemistry/ Immunofluorescence

Detection of Mouse GLI-3 by Immunocytochemistry/ Immunofluorescence

Analysis of Ptch receptor-dependent inhibition of Hh signaling. (A,C,E) Immunocytochemistry detection of Gli3 (A), Gli1 (C) and Gli2 (E) in the wild-type, Ptch1−/−, and Ptch2−/− MEFs. Scale bar 50 μm. (B,D,F) Quantification of the intensity of the nuclear GLI3 (B), GLI1 (D), and GLI2 (F) in wild-type, Ptch1−/−, and Ptch2−/− MEFs. **p ≤ 0.005, ***p ≤ 0.0005, ****p < 0.0001. (G) Western blot showing nuclear (N) and cellular (C) expression of Gli2 (FL = full length and RF = repressor form) in wild-type, Ptch1−/− and Ptch2−/− MEFs. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/35574464), licensed under a CC-BY license. Not internally tested by R&D Systems.

Applications for GLI-3 Antibody

Application
Recommended Usage

Chromatin Immunoprecipitation (ChIP)

5 µg/106 cells
Sample: PMA and ionomycin treated Jurkat human acute T cell leukemia cell line chromatin, GLI-1 promoter detected by standard PCR

Immunocytochemistry

5-15 µg/mL
Sample: Immersion fixed HeLa human cervical epithelial carcinoma cell line

Western Blot

1 µg/mL
Sample: Mouse embryo tissue

Reviewed Applications

Read 5 reviews rated 4.8 using AF3690 in the following applications:

Formulation, Preparation, and Storage

Purification

Antigen Affinity-purified

Reconstitution

Reconstitute at 0.2 mg/mL in sterile PBS. For liquid material, refer to CoA for concentration.

Formulation

Lyophilized from a 0.2 μm filtered solution in PBS with Trehalose. *Small pack size (SP) is supplied either lyophilized or as a 0.2 µm filtered solution in PBS.

Shipping

Lyophilized product is shipped at ambient temperature. Liquid small pack size (-SP) is shipped with polar packs. Upon receipt, store immediately at the temperature recommended below.

Stability & Storage

Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
  • 12 months from date of receipt, -20 to -70 °C as supplied.
  • 1 month, 2 to 8 °C under sterile conditions after reconstitution.
  • 6 months, -20 to -70 °C under sterile conditions after reconstitution.

Calculators

The reconstitution calculator allows you to quickly calculate the volume of a reagent to reconstitute your vial. Simply enter the mass of reagent and the target concentration and the calculator will determine the rest.

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Background: GLI-3

Human GLI-3 is a 190 kDa transcription factor of the C2H2-type zinc-finger protein family. GLI-3 appears to play a role in limb and brain development, probably through sonic hedgehog signaling. GLI-3 is regulated by PKA-mediated phosphorylation. Unphosphorylated full-length GLI-3 is a transcriptional activator, while phosphorylated GLI-3 is cleaved after amino acid 700 to generate an N-terminal 83 kDa transcriptional repressor. Over the region used as immunogen, human GLI-3 is 95% identical to the corresponding mouse and canine protein sequences.

Long Name

GLI-Kruppel family member GLI3

Alternate Names

ACLS, ADD, Bph, GCPS, GLI3, PAPA1, PAPB, Pdn, PHS, PPDIV, Xt

Entrez Gene IDs

2737 (Human); 14634 (Mouse); 140588 (Rat)

Gene Symbol

GLI3

UniProt

P10071

Additional GLI-3 Products

Product Documents for GLI-3 Antibody

Certificate of Analysis

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Product Specific Notices for GLI-3 Antibody

For research use only

Customer Reviews for GLI-3 Antibody (5)

4.8 out of 5
5 Customer Ratings
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4 Stars
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Customer Images

Showing  1 - 5 的 5 reviews Showing All
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  • Human/Mouse GLI-3 Antibody
    Name: Anonymous
    Application: Western Blot
    Sample Tested: Cell Lysates
    Species: Mouse
    Verified Customer | Posted 05/22/2019
    GLI-3 Antibody AF3690
  • Name: Anonymous
    Application: Western Blot
    Sample Tested: NIH-3T3 cells
    Species: Mouse
    Verified Customer | Posted 11/13/2015
    Gli3 Western Blot
    GLI-3 Antibody AF3690
  • Human/Mouse GLI‑3 Antibody
    Name: Anonymous
    Application: Western Blot
    Sample Tested: Mouse embryo cell lysate
    Species: Mouse
    Verified Customer | Posted 10/26/2015
    Specificity: Specific<br />Sensitivity: Sensitive<br />Buffer: TBST<br />Dilution: 1/1000
    GLI-3 Antibody AF3690
  • Name: Anonymous
    Application: Western Blot
    Sample Tested: See PMID 22173325
    Species: Mouse
    Verified Customer | Posted 01/08/2015
  • Name: Anonymous
    Application: Western Blot
    Sample Tested: See PMID 22547067
    Species: Mouse
    Verified Customer | Posted 01/08/2015

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Protocols

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Associated Pathways

TGF-beta Signaling Pathways
TGF-beta Signaling Pathway Thumbnail