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47193
POMC (AH26) Mouse mAb
Primary Antibodies
Monoclonal Antibody

POMC (AH26) Mouse mAb #47193

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  1. IHC
Immunohistochemistry Image 1: POMC (AH26) Mouse mAb
Immunohistochemical analysis of paraffin-embedded normal human pituitary gland using POMC (AH26) Mouse mAb.
Immunohistochemistry Image 2: POMC (AH26) Mouse mAb
Immunohistochemical analysis of paraffin-embedded normal human pituitary gland (left-top) and various paraffin-embedded human tissues: normal colon (middle-top), normal liver (right-top), urothelial carcinoma (left-bottom), non-small cell lung carcinoma (middle-bottom), and ductal breast carcinoma (right-bottom) using POMC (AH26) Mouse mAb. Note the lack of staining in tissues other than the pituitary gland.
Immunohistochemistry Image 3: POMC (AH26) Mouse mAb
Immunohistochemical analysis of paraffin-embedded normal human pituitary gland using POMC (AH26) Mouse mAb (left) compared to concentration-matched Mouse (G3A1) mAb IgG1 Isotype Control #5415 (right). 
To Purchase # 47193S
Product # Size Price
47193S
100 µl $ 285

Supporting Data

REACTIVITY H
SENSITIVITY Endogenous
MW (kDa)
Source/Isotype Mouse IgG1 kappa

Application Key:

  • WB-Western Blot
  • IP-Immunoprecipitation
  • IHC-Immunohistochemistry
  • ChIP-Chromatin Immunoprecipitation
  • IF-Immunofluorescence
  • F-Flow Cytometry
  • E-P-ELISA-Peptide

Species Cross-Reactivity Key:

  • H-Human
  • M-Mouse
  • R-Rat
  • Hm-Hamster
  • Mk-Monkey
  • Vir-Virus
  • Mi-Mink
  • C-Chicken
  • Dm-D. melanogaster
  • X-Xenopus
  • Z-Zebrafish
  • B-Bovine
  • Dg-Dog
  • Pg-Pig
  • Sc-S. cerevisiae
  • Ce-C. elegans
  • Hr-Horse
  • All-All Species Expected

Product Usage Information

Application Dilution
Immunohistochemistry (Paraffin) 1:800 - 1:3200

Storage

Supplied in 10 mM PBS containing 0.05% BSA and 0.05% sodium azide. Stable for 24 months when stored at -20°C. This product will freeze at -20°C so it is recommended to aliquot into single-use vials to avoid multiple freeze/thaw cycles. A slight precipitate may be present, but will not interfere with antibody performance.

Protocol

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Immunohistochemistry (Paraffin)

A. Solutions and Reagents

NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalent grade water.

  1. Xylene.
  2. Ethanol, anhydrous denatured, histological grade (100% and 95%).
  3. Deionized water (dH2O).
  4. Hematoxylin (optional).
  5. Wash Buffer:
    1. 1X Tris Buffered Saline with Tween® 20 (TBST): To prepare 1L 1X TBST add 100 ml 10X Tris Buffered Saline with Tween ® 20 (#9997) to 900 ml dH 2O, mix.
  6. SignalStain® Antibody Diluent (#8112).
  7. 1X Citrate Unmasking Solution: To prepare 250 mL of 1X citrate unmasking solution, dilute 25 ml of SignalStain® Citrate Unmasking Solution (10X) (#14746) with 225 mL of dH2O.
  8. 3% Hydrogen Peroxide: To prepare 100 ml, add 10 ml 30% H 2O2 to 90 ml dH2O.
  9. Blocking Solution: TBST/5% Normal Goat Serum or 1X Animal-Free Blocking Solution.
    1. TBST/5% Normal Goat Serum: to 5 ml 1X TBST, add 250 µl Normal Goat Serum (#5425).
    2. 1X Animal-Free Blocking Solution: to 4 mL of dH2O add 1 ml of Animal-Free Blocking Solution (5X) (#15019).
  10. Detection System: SignalStain® Boost IHC Detection Reagents (HRP, Mouse #8125).
  11. Substrate: SignalStain® DAB Substrate Kit (#8059).
  12. Hematoxylin: Hematoxylin (#14166).
  13. Mounting Medium: SignalStain® Mounting Medium (#14177).

B. Deparaffinization/Rehydration

NOTE: Do not allow slides to dry at any time during this procedure.

  1. Deparaffinize/hydrate sections:
    1. Incubate sections in three washes of xylene for 5 min each.
    2. Incubate sections in two washes of 100% ethanol for 10 min each.
    3. Incubate sections in two washes of 95% ethanol for 10 min each.
  2. Wash sections two times in dH2O for 5 min each.

C. Antigen Unmasking

For Citrate: Heat slides in a microwave submersed in 1X citrate unmasking solution until boiling is initiated; follow with 10 min at a sub-boiling temperature (95°-98°C). Cool slides on bench top for 30 min.

D. Staining

  1. Wash sections in dH2O three times for 5 min each.
  2. Incubate sections in 3% hydrogen peroxide for 10 min.
  3. Wash sections in dH2O two times for 5 min each.
  4. Wash sections in wash buffer for 5 min.
  5. Block each section with 100–400 µl of preferred blocking solution for 1 hr at room temperature.
  6. Remove blocking solution and add 100–400 µl primary antibody diluted in SignalStain® Antibody Diluent (#8112) to each section. Incubate overnight at 4°C.
  7. Equilibrate SignalStain® Boost Detection Reagent (HRP, Mouse #8125) to room temperature.
  8. Remove antibody solution and wash sections with wash buffer three times for 5 min each.
  9. Cover section with 1–3 drops SignalStain® Boost Detection Reagent (HRP, Mouse #8125) as needed. Incubate in a humidified chamber for 30 min at room temperature.
  10. Wash sections three times with wash buffer for 5 min each.
  11. Add 1 drop (30 µl) SignalStain® DAB Chromogen Concentrate to 1 ml SignalStain® DAB Diluent and mix well before use.
  12. Apply 100–400 µl SignalStain® DAB to each section and monitor closely. 5–10 min generally provides an acceptable staining intensity.
  13. Immerse slides in dH2O.
  14. If desired, counterstain sections with hematoxylin (#14166).
  15. Wash sections in dH2O two times for 5 min each.
  16. Dehydrate sections:
    1. Incubate sections in 95% ethanol two times for 10 sec each.
    2. Repeat in 100% ethanol, incubating sections two times for 10 sec each.
    3. Repeat in xylene, incubating sections two times for 10 sec each.
  17. Mount sections with coverslips and mounting medium (#14177).
DETECTION REAGENT/SUBSTRATE COMPATIBILITY
RECOMMENDED
DETECTION REAGENTS
SignalStain® Boost IHC Detection Reagent (HRP, Mouse) #8125 SignalStain® Boost IHC Detection Reagent (AP, Mouse) #31926
COMPATIBLE
CHROMOGEN
SignalStain® DAB Substrate Kit #8059 SignalStain® Vibrant Red Alkaline Phosphatase Substrate Kit #76713
SignalStain® Vivid Purple Peroxidase Substrate Kit #96632  

NOTE: Use of detection reagents other than those specified in this protocol may require further optimization of the primary antibody to account for the different sensitivities of the detection reagents.

posted February 2010

revised April 2020

Protocol Id: 1991

Specificity / Sensitivity

POMC (AH26) Mouse mAb recognizes endogenous levels of total POMC protein.

Species Reactivity:

Human

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to amino acids 1-24 of human POMC protein.

Background

Pro-opiomelanocortin (POMC) is a precursor protein expressed in the pituitary and the brain where it is processed into several peptide hormones and neuropeptides. Among these peptides are ACTH, α- and β-MSH, β-and γ-LPH, CLIP, β-endorphin, and N-POMC (1). POMC is involved in hypothalamic circuits regulating feeding behavior and POMC-producing neurons promote satiety (2). POMC neurons are also the target of leptin and insulin for the promotion of the browning of white fat (3).
  1. Chretien, M. and Mbikay, M. (2016) J Mol Endocrinol, T49-62. doi: 10.1530/JME-15-0261.
  2. Yang, Y. et al. (2011) Cell 146, 992-1003.
  3. Dodd, G.T. et al. (2015) Cell 160, 88-104.

Pathways & Proteins

Explore pathways + proteins related to this product.

Limited Uses

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For Research Use Only. Not For Use In Diagnostic Procedures.
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