Detection of Phospho-GSK‑3 alpha / beta (S21/S9) in Jurkat Human Cell Line by Flow Cytometry.
Jurkat human acute T cell leukemia cells, untreated (open histogram, yellow line), or treated with PI3-kinase inhibitors (dark orange filled histogram), were stained with Human/Mouse/Rat Phospho-GSK‑3 alpha / beta (S21/S9) Antigen Affinity‑purified Polyclonal Antibody (Catalog # AF1590) or control antibody (Catalog # AB-105-C, blue open histogram), followed by Phycoerythrin-conjugated Anti-Rabbit IgG Secondary Antibody (Catalog # F0110). To facilitate intracellular staining, cells were fixed with paraformaldehyde and permeabilized with saponin.
Detection of Human Phospho-GSK‑3 alpha (S21) and GSK-3 beta (S9) by Western Blot.
Western blot shows lysates of HeLa human cervical epithelial carcinoma cell line untreated (-) or treated (+) with 200 nM PMA for 20 minutes and MCF‑7 human breast cancer cell line untreated or treated with 100 ng/mL Recombinant Human IGF-1 (Catalog # 291‑G1) for 15 minutes. PVDF membrane was probed with 0.2 µg/mL of Human/Mouse/Rat Phospho-GSK‑3 alpha / beta (S21/S9) Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1590), followed by HRP-conjugated Anti-Rabbit IgG Secondary Antibody (Catalog # HAF008). Specific bands were detected for Phospho-GSK‑3 alpha (S21) and GSK-3 beta (S9) at approximately 51 and 46 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.
Preparation and Storage
Reconstitute at 0.2 mg/mL in sterile PBS.
Reconstitution Buffer Available
The product is shipped at ambient temperature. Upon receipt, store it immediately at the temperature recommended below. *Small pack size (SP) is shipped with polar packs. Upon receipt, store it immediately at -20 to -70 °C
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.
Background: GSK-3 alpha/beta
GSK-3 is a Ser/Thr kinase first identified as an inactivator of Glycogen Synthase. GSK-3 acts as a multifunctional downstream switch that determines the output of numerous signaling pathways. There are two mammalian GSK-3 isoforms encoded by distinct genes, GSK-3 alpha and GSK-3 beta, which are structurally similar, but functionally non-identical. GSK-3a is inhibited by phosphorylation at S21 by Akt and other kinases. GSK-3 alpha and GSK-3 beta share 85% amino acid identity. Dysregulated GSK-3 has been implicated in several diseases including type II diabetes, Alzheimer's disease, bipolar disorder, and cancer.
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