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Purified Mouse Anti-Glucagon
Purified Mouse Anti-Glucagon
Flow cytometric analysis of Glucagon expression in human Glucagon-transfected 293F cells and a mouse pancreatic α cell line. LEFT PANEL: Untransfected (dashed line histogram) and human Glucagon-transfected (solid line histogram) 293F cells were fixed with BD Cytofix™ Fixation Buffer (Cat. No. 554655) and permeabilized with BD Phosflow™ Perm Buffer III (Cat. No. 558050). The cells were washed and then stained with Purified Mouse Anti-Glucagon (Cat. No. 565859) followed by APC Goat Anti-Mouse Ig (Cat. No. 550826). RIGHT PANEL: Alpha TC1-6 cells (ATCC CRL-2934) were fixed with BD Cytofix™ Fixation Buffer (Cat. No. 554655) and permeabilized with BD Phosflow™ Perm Buffer III (Cat. No. 558050). The cells were washed and then stained with either Purified Mouse IgG1, κ isotype control (Cat. No. 554121, dashed-line histogram) or Purified Mouse Anti-Glucagon antibody (Cat. No. 565859, solid-line histogram) followed by APC Goat Anti-Mouse Ig (Cat. No. 550826). All fluorescence histograms were derived from gated events with the forward and side light-scatter characteristics of intact cells. Flow cytometry was performed on a BD FACSCanto™ II flow cytometry system.
Purified Mouse Anti-Glucagon

LEFT PANEL: Western blot analysis of Glucagon expression. Lysate from human Glucagon-transfected 293F cells was prepared for electrophoresis (SDS-PAGE) in a 2D Tris-Glycine polyacrylamide gel. The proteins were transferred to PVDF membranes and then probed with 4 (lane 1), 2 (lane 2), and 1 (lane 3) µg/mL of Purified Mouse Anti-Glucagon (Cat. No. 565859). Specific staining was detected with HRP Goat Anti-Mouse Ig (Cat No 554002).

RIGHT PANEL: Immunohistochemical staining of Glucagon in human, rat, and mouse islets of Langerhans. Following antigen retrieval with BD Retrievagen A Buffer (Cat. No. 550524), sections from formalin-fixed, paraffin-embedded human (top row), rat (middle row), and mouse (bottom row) pancreata were blocked using an Avidin/Biotin Blocking Kit (Vector Laboratories, Cat. No. SP-2001) as recommended by the manufacturer. The sections were then stained overnight with either Purified Mouse IgG1 κ Isotype Control (Cat. No. 550878, left column) or Purified Mouse Anti-Glucagon (Cat. No. 565859, right column). A three-step staining procedure that employs either Biotin Goat Anti-Mouse Ig (Cat. No. 550337, top and middle rows) or Biotin Rat Anti-Mouse IgG1 (Cat. No.553441, bottom row), Streptavidin HRP (Cat. No. 550946) and DAB (Cat. No. 550880) was used to reveal the primary staining reagents. Counterstaining was with Hematoxylin. The typical localization of the α cells is seen: distributed throughout human islets and at the periphery in rodent islets. Original magnifications: 40X.

Flow cytometric analysis of Glucagon expression in human Glucagon-transfected 293F cells and a mouse pancreatic α cell line. LEFT PANEL: Untransfected (dashed line histogram) and human Glucagon-transfected (solid line histogram) 293F cells were fixed with BD Cytofix™ Fixation Buffer (Cat. No. 554655) and permeabilized with BD Phosflow™ Perm Buffer III (Cat. No. 558050). The cells were washed and then stained with Purified Mouse Anti-Glucagon (Cat. No. 565859) followed by APC Goat Anti-Mouse Ig (Cat. No. 550826). RIGHT PANEL: Alpha TC1-6 cells (ATCC CRL-2934) were fixed with BD Cytofix™ Fixation Buffer (Cat. No. 554655) and permeabilized with BD Phosflow™ Perm Buffer III (Cat. No. 558050). The cells were washed and then stained with either Purified Mouse IgG1, κ isotype control (Cat. No. 554121, dashed-line histogram) or Purified Mouse Anti-Glucagon antibody (Cat. No. 565859, solid-line histogram) followed by APC Goat Anti-Mouse Ig (Cat. No. 550826). All fluorescence histograms were derived from gated events with the forward and side light-scatter characteristics of intact cells. Flow cytometry was performed on a BD FACSCanto™ II flow cytometry system.

LEFT PANEL: Western blot analysis of Glucagon expression. Lysate from human Glucagon-transfected 293F cells was prepared for electrophoresis (SDS-PAGE) in a 2D Tris-Glycine polyacrylamide gel. The proteins were transferred to PVDF membranes and then probed with 4 (lane 1), 2 (lane 2), and 1 (lane 3) µg/mL of Purified Mouse Anti-Glucagon (Cat. No. 565859). Specific staining was detected with HRP Goat Anti-Mouse Ig (Cat No 554002).

RIGHT PANEL: Immunohistochemical staining of Glucagon in human, rat, and mouse islets of Langerhans. Following antigen retrieval with BD Retrievagen A Buffer (Cat. No. 550524), sections from formalin-fixed, paraffin-embedded human (top row), rat (middle row), and mouse (bottom row) pancreata were blocked using an Avidin/Biotin Blocking Kit (Vector Laboratories, Cat. No. SP-2001) as recommended by the manufacturer. The sections were then stained overnight with either Purified Mouse IgG1 κ Isotype Control (Cat. No. 550878, left column) or Purified Mouse Anti-Glucagon (Cat. No. 565859, right column). A three-step staining procedure that employs either Biotin Goat Anti-Mouse Ig (Cat. No. 550337, top and middle rows) or Biotin Rat Anti-Mouse IgG1 (Cat. No.553441, bottom row), Streptavidin HRP (Cat. No. 550946) and DAB (Cat. No. 550880) was used to reveal the primary staining reagents. Counterstaining was with Hematoxylin. The typical localization of the α cells is seen: distributed throughout human islets and at the periphery in rodent islets. Original magnifications: 40X.

Product Details
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BD Pharmingen™
GLP1; GLP2; GRPP; GCG
Human (QC Testing), Mouse, Rat (Tested in Development)
Mouse BALB/c IgG1, κ
Human glucagon Recombinant Protein
Intracellular staining (flow cytometry) (Routinely Tested), Immunofluorescence, Immunohistochemistry-formalin (antigen retrieval required), Western blot (Tested During Development)
0.5 mg/ml
AB_2739381
Aqueous buffered solution containing ≤0.09% sodium azide.
RUO


Preparation And Storage

Store undiluted at 4°C. The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography.

Product Notices

  1. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
  2. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
  3. Caution: Sodium azide yields highly toxic hydrazoic acid under acidic conditions. Dilute azide compounds in running water before discarding to avoid accumulation of potentially explosive deposits in plumbing.
  4. Sodium azide is a reversible inhibitor of oxidative metabolism; therefore, antibody preparations containing this preservative agent must not be used in cell cultures nor injected into animals. Sodium azide may be removed by washing stained cells or plate-bound antibody or dialyzing soluble antibody in sodium azide-free buffer. Since endotoxin may also affect the results of functional studies, we recommend the NA/LE (No Azide/Low Endotoxin) antibody format, if available, for in vitro and in vivo use.
  5. Species cross-reactivity detected in product development may not have been confirmed on every format and/or application.
  6. An isotype control should be used at the same concentration as the antibody of interest.
565859 Rev. 1
Antibody Details
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U16-850

The U16-850 monoclonal antibody specifically binds to glucagon, a member of the secretin family of active peptides. Glucagon is an evolutionarily conserved peptide hormone that participates in the regulation of carbohydrate metabolism by counteracting the effects of insulin. Glucagon is produced by α cells in the islets of Langerhans of the pancreas. The CGC gene encodes the precursor molecule preproglucagon, which is cleaved to form proglucagon that is in turn cleaved to form at least four distinct peptides, including glucagon, in different tissues. Hypoglycemia causes the secretion of glucagon, which binds to the class B G-protein-coupled glucagon receptor that is mainly expressed in liver and kidney, causing reduced glycogenesis and glycolysis and increased glycogenolysis and gluconeogenesis. The expression of glucagon can be used to monitor the pancreatic differentiation of pluripotent stem cells.

565859 Rev. 1
Format Details
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Purified
Tissue culture supernatant is purified by either protein A/G or affinity purification methods. Both methods yield antibody in solution that is free of most other soluble proteins, lipids, etc. This format provides pure antibody that is suitable for a number of downstream applications including: secondary labeling for flow cytometry or microscopy, ELISA, Western blot, etc.
Purified
565859 Rev.1
Citations & References
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Development References (6)

  1. Bowerman M, Michalski JP, Beauvais A, Murray LM, DeRepentigny Y, Kothary R. Defects in pancreatic development and glucose metabolism in SMN-depleted mice independent of canonical spinal muscular atrophy neuromuscular pathology.. Hum Mol Genet. 2014; 23(13):3432-44. (Biology). View Reference
  2. D'Amour KA, Bang AG, Eliazer S, et al . Production of pancreatic hormone-expressing endocrine cells from human embryonic stem cells. Nat Biotechnol. 2006; 24(12):1481-1483. (Biology). View Reference
  3. Jiang G, Zhang BB. Glucagon and regulation of glucose metabolism.. Am J Physiol Endocrinol Metab. 2003; 284(4):E671-8. (Biology). View Reference
  4. Kelly OG, Chan MY, Martinson LA, et al. Cell-surface markers for the isolation of pancreatic cell types derived from human embryonic stem cells. Nat Biotechnol. 2011; 29(8):750-756. (Biology). View Reference
  5. Thomsen J, Kristiansen K, Brunfeldt K, Sundby F. The amino acid sequence of human glucagon.. FEBS Lett. 1972; 21(3):315-319. (Biology). View Reference
  6. Witt S, Dietz H, Ziegler B, Keilacker H, Ziegler M. Production and use of monoclonal glucagon and insulin antibodies--reduction of pancreatic insulin in rats by treatment with complete Freund's adjuvant. Acta Histochem Suppl. 1988; 35:217-223. (Biology). View Reference
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565859 Rev. 1

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Comparisons, where applicable, are made against older BD Technology, manual methods or are general performance claims.  Comparisons are not made against non-BD technologies, unless otherwise noted.

For Research Use Only. Not for use in diagnostic or therapeutic procedures.