Revision 1

#9369Store at -20C

1 Kit

(5 x 20 microliters)

Cell Signaling Technology

Orders: 877-616-CELL (2355) [email protected]

Support: 877-678-TECH (8324)

Web: [email protected] cellsignal.com

3 Trask LaneDanversMassachusetts01923USA
For Research Use Only. Not for Use in Diagnostic Procedures.
Product Includes Product # Quantity Mol. Wt Isotype/Source
GSK-3α (D80E6) Rabbit mAb 4337 20 µl 51 kDa Rabbit IgG
Phospho-GSK-3α (Ser21) (36E9) Rabbit mAb 9316 20 µl 51 kDa Rabbit 
Phospho-GSK-3β (Ser9) (D85E12) XP® Rabbit mAb 5558 20 µl 46 kDa Rabbit IgG
Phospho-GSK-3α/β (Ser21/9) (D17D2) Rabbit mAb 8566 20 µl 46 GSK-3beta, 51 GSK-3alpha kDa Rabbit IgG
GSK-3β (D5C5Z) XP® Rabbit mAb 12456 20 µl 46 kDa Rabbit IgG
Anti-rabbit IgG, HRP-linked Antibody 7074 100 µl Goat 

Please visit cellsignal.com for individual component applications, species cross-reactivity, dilutions, protocols, and additional product information.

Description

The GSK-3 Antibody Sampler Kit contains primary and secondary antibodies to perform two Western blots with each antibody.

Storage

Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.

Background

Glycogen synthase kinase-3 (GSK-3) was initially identified as an enzyme that regulates glycogen synthesis in response to insulin (1). GSK-3 is a ubiquitously expressed serine/threonine protein kinase that phosphorylates and inactivates glycogen synthase. GSK-3 is a critical downstream element of the PI3K/Akt cell survival pathway whose activity can be inhibited by Akt-mediated phosphorylation at Ser21 of GSK-3α and Ser9 of GSK-3β (2,3). GSK-3 has been implicated in the regulation of cell fate in Dictyostelium and is a component of the Wnt signaling pathway required for Drosophila, Xenopus, and mammalian development (4). GSK-3 has been shown to regulate cyclin D1 proteolysis and subcellular localization (5).

  1. Welsh, G.I. et al. (1996) Trends Cell Biol 6, 274-9.
  2. Srivastava, A.K. and Pandey, S.K. (1998) Mol Cell Biochem 182, 135-41.
  3. Cross, D.A. et al. (1995) Nature 378, 785-9.
  4. Nusse, R. (1997) Cell 89, 321-3.
  5. Diehl, J.A. et al. (1998) Genes Dev 12, 3499-511.

Background References

    Trademarks and Patents

    Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
    All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

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    Revision 1
    #9369

    GSK-3 Antibody Sampler Kit

    GSK-3 Antibody Sampler Kit: Image 1 Expand Image
    Simple Western™ analysis of lysates (0.1 mg/mL) from PDGF-treated 3T3 cells using Phospho-GSK-3β (Ser9) (D85E12) XP® Rabbit mAb #5558. The virtual lane view (left) shows the target band (as indicated) at 1:50 and 1:250 dilutions of primary antibody. The corresponding electropherogram view (right) plots chemiluminescence by molecular weight along the capillary at 1:50 (blue line) and 1:250 (green line) dilutions of primary antibody. This experiment was performed under reducing conditions on the Jess™ Simple Western instrument from ProteinSimple, a BioTechne brand, using the 12-230 kDa separation module.
    GSK-3 Antibody Sampler Kit: Image 2 Expand Image
    Western blot analysis of extracts from wild-type, GSK-3α (-/-), and GSK3β (-/-) mouse embryonic fibroblasts (MEFs) using GSK-3β (D5C5Z) XP® Rabbit mAb (upper) and GSK-3α/β (D75D3) XP® Rabbit mAb #5676 (lower). (MEF wild type, GSK-3α (-/-), and GSK-3β (-/-) cells were kindly provided by Dr. Jim Woodgett, University of Toronto, Canada).
    GSK-3 Antibody Sampler Kit: Image 3 Expand Image
    Western blot analysis of extracts from various cell types using GSK-3α (D80E6) Rabbit mAb.
    GSK-3 Antibody Sampler Kit: Image 4 Expand Image
    Western blot analysis of extracts from GSK-3α (-/-) (lanes 1,2) and GSK-3β (-/-) (lanes 3,4) mouse embryonic fibroblast (MEF) cells, λ phosphatase or PDGF-treated, using Phospho-GSK-3β (Ser9) (D85E12) XP® Rabbit mAb (upper) and α/β-Tubulin Antibody #2148 (lower). (MEF wild type, GSK-3α (-/-) and GSK-3β (-/-) cells were kindly provided by Dr. Jim Woodgett, University of Toronto, Canada).
    GSK-3 Antibody Sampler Kit: Image 5 Expand Image
    After the primary antibody is bound to the target protein, a complex with HRP-linked secondary antibody is formed. The LumiGLO® is added and emits light during enzyme catalyzed decomposition.
    GSK-3 Antibody Sampler Kit: Image 6 Expand Image
    Western blot analysis of extracts from COS-7 cells, λ phosphatase- or PDGF-treated (100 μg/ml, 15 min), and PC-3 cells, untreated or λ phosphatase-treated, using Phospho-GSK-3α/β (Ser21/9) (D17D2) Rabbit mAb (upper) and GSK-3α/β (D75D3) XP® Rabbit mAb #5676 (lower).
    GSK-3 Antibody Sampler Kit: Image 7 Expand Image
    Western blot analysis of extracts from COS-7 cells, λ-phosphatase or PDGF-treated, using Phospho-GSK-3α (Ser21) (36E9) Rabbit mAb (upper) or GSK-3α Antibody #9338 (lower).
    GSK-3 Antibody Sampler Kit: Image 8 Expand Image
    Immunoprecipitation of GSK-3β from PC-12 cell extracts, using Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (lane 2) or GSK-3β (D5C5Z) XP® Rabbit mAb (lane 3). Lane 1 is 10% input. Western blot analysis was performed using GSK-3β (D5C5Z) XP® Rabbit mAb.
    GSK-3 Antibody Sampler Kit: Image 9 Expand Image
    Western blot analysis of extracts from PC-3 cells, untreated or LY294002/wortmannin-treated, using Phospho-GSK-3β (Ser9) (D85E12) XP® Rabbit mAb (upper) or GSK-3β (27C10) Rabbit mAb #9315 (lower).
    GSK-3 Antibody Sampler Kit: Image 10 Expand Image
    Western blot analysis of extracts from GSK-3α (-/-) and GSK-3β (-/-) mouse embryonic fibroblasts (MEF), untreated or insulin-treated (100 ng/ml, 20 min) , using Phospho-GSK-3α/β (Ser21/9) (D17D2) Rabbit mAb (upper) and GSK-3α/β (D75D3) XP® Rabbit mAb #5676 (lower). (MEF GSK-3α (-/-) and GSK-3β (-/-) cells were kindly provided by Dr. Jim Woodgett, University of Toronto, Canada).
    GSK-3 Antibody Sampler Kit: Image 11 Expand Image
    Immunohistochemical analysis of paraffin-embedded human breast carcinoma , untreated (left) or lambda phosphatase treated (right), using Phospho-GSK-3alpha (Ser 21) (36E9) Rabbit mAb.
    GSK-3 Antibody Sampler Kit: Image 12 Expand Image
    Immunohistochemical analysis of paraffin-embedded human breast carcinoma using GSK-3β (D5C5Z) XP® Rabbit mAb.
    GSK-3 Antibody Sampler Kit: Image 13 Expand Image
    Confocal immunofluorescent analysis of wild type mouse embryonic fibroblasts (MEFs) (top row), GSK-3β (-/-) MEFs (middle row) , or PC-3 cells (bottom row), untreated (left), LY294002- and Wortmannin-treated (#9901 and #9951 respectively; center) or lambda phosphatase-treated (right), using Phospho-GSK-3β (Ser9) (D85E12) XP® Rabbit mAb (green). Actin filaments were labeled with DY-554 phalloidin (red). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye). (MEF wild type and GSK-3β (-/-) cells were kindly provided by Dr. Jim Woodgett, University of Toronto, Canada).
    GSK-3 Antibody Sampler Kit: Image 14 Expand Image
    Immunohistochemical analysis of paraffin-embedded LNCaP cells, untreated (left) or LY294002-treated (right), using Phospho-GSK-3alpha (Ser 21) (36E9) Rabbit mAb.
    GSK-3 Antibody Sampler Kit: Image 15 Expand Image
    Immunohistochemical analysis of paraffin-embedded mouse pancreas using GSK-3β (D5C5Z) XP® Rabbit mAb.
    GSK-3 Antibody Sampler Kit: Image 16 Expand Image
    Flow cytometric analysis of NIH/3T3 cells, untreated (blue) or PDGF-treated (green), using Phospho-GSK-3β (Ser9) (D85E12) XP® Rabbit mAb.
    GSK-3 Antibody Sampler Kit: Image 17 Expand Image
    Immunohistochemical analysis of paraffin-embedded human breast carcinoma using Phospho-GSK-3α (Ser21) (36E9) Rabbit mAb in the presence of control peptide (left) or Phospho-GSK-3α (Ser21) (36E9) Blocking Peptide #1027 (right).
    GSK-3 Antibody Sampler Kit: Image 18 Expand Image
    Immunohistochemical analysis of paraffin-embedded MEF cell pellets, wild type (left), GSK-3α (-/-) (middle) and GSK-3β (-/-) (right) using GSK-3β (D5C5Z) XP® Rabbit mAb. (MEF wild type, GSK-3β (-/-), and GSK-3α (-/-) cells were kindly provided by Dr. Jim Woodgett, University of Toronto, Canada).
    GSK-3 Antibody Sampler Kit: Image 19 Expand Image
    Immunohistochemical analysis of paraffin-embedded human lung carcinoma using Phospho-GSK-3alpha (Ser 21) (36E9) Rabbit mAb.
    GSK-3 Antibody Sampler Kit: Image 20 Expand Image
    Confocal immunofluorescent analysis of wild-type mouse embryonic fibroblasts (MEFs) (left), GSK-3α (-/-) MEFs (center)and GSK-3β (-/-) MEFs (right) using GSK-3β (D5C5Z) XP® Rabbit mAb (green). Actin filaments were labeled with DY-554 phalloidin (red). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye). (MEF wild type, GSK-3α (-/-), and GSK-3β (-/-) cells were kindly provided by Dr. Jim Woodgett, University of Toronto, Canada).
    GSK-3 Antibody Sampler Kit: Image 21 Expand Image
    Flow cytometric analysis of GSK-3β (-/-) MEFs (blue, negative) and wild type mouse embryonic fibroblasts (MEFs) (green, positive) using GSK-3β (D5C5Z) XP® Rabbit mAb or a concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (dashed lines). Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody. (MEF wild type and GSK-3β (-/-) cells were kindly provided by Dr. Jim Woodgett, University of Toronto, Canada).