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13888
Galectin-1/LGALS1 (D608T) Rabbit mAb (IHC Formulated)

Galectin-1/LGALS1 (D608T) Rabbit mAb (IHC Formulated) #13888

APPLICATIONS

REACTIVITY SENSITIVITY MW (kDa) Isotype
H M R Endogenous Rabbit IgG
IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded human breast carcinoma using Galectin-1/LGALS1 (D608T) Rabbit mAb (IHC Formulated) in the presence of control peptide (left) or antigen specific peptide (right).

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IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded human ovarian mucinous cystadenocarcinoma using Galectin-1/LGALS1 (D608T) Rabbit mAb (IHC Formulated).

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IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded A549 (left) and LNCaP (right) cell pellets using Galectin-1/LGALS1 (D608T) Rabbit mAb (IHC Formulated).

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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 dH20, 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% H2O2 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, Rabbit #8114).
  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, Rabbit #8114) 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, Rabbit #8114) 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. 1–10 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).

posted February 2010

revised March 2016

Protocol Id: 283

Application Dilutions
Immunohistochemistry (Paraffin) 1:250
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.

Galectin-1/LGALS1 (D608T) Rabbit mAb (IHC Formulated) recognizes endogenous levels of total galectin-1/LGALS1 protein.

Species Reactivity:

Human, Mouse, Rat

Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues near the amino terminus of human galectin-1/LGALS1 protein.

Galectins are a family of β-galactose binding proteins that are characterized by their affinity for poly-N-acetyllactosamine-enriched glycoconjugates and their carbohydrate-binding site (1,2). Members of the galectin family have been implicated in a variety of biological functions including cell adhesion (3), growth regulation (4), cytokine production (5), T cell apoptosis (6), and immune responses (7). Galectin-1/LGALS1 has been shown to be expressed in a wide range of tissues and cell types. The level and pattern of expression of galectin-1 have been shown to change during development (8). In addition to a role in developmental processes, galectin-1 has been shown to be involved in central immune tolerance and may function in tumorigenesis by modulating the immune response to the tumor (9,10). Research studies have shown that galectin-1 expression is increased in several human cancers, suggesting a correlation with metastatic potential (10).

  1. Barondes, S.H. et al. (1994) Cell 76, 597-8.
  2. Barondes, S.H. et al. (1994) J Biol Chem 269, 20807-10.
  3. Offner, H. et al. (1990) J Neuroimmunol 28, 177-84.
  4. Wells, V. and Mallucci, L. (1991) Cell 64, 91-7.
  5. Filer, A. et al. (2009) Arthritis Rheum 60, 1604-14.
  6. Perillo, N.L. et al. (1995) Nature 378, 736-9.
  7. Cooper, D.N. et al. (1991) J Cell Biol 115, 1437-48.
  8. Puche, A.C. et al. (1996) Dev Biol 179, 274-87.
  9. van den Brûle, F. et al. (2003) Lab Invest 83, 377-86.
  10. Salatino, M. et al. (2008) Expert Opin Biol Ther 8, 45-57.
Entrez-Gene Id
3956
Swiss-Prot Acc.
P09382
For Research Use Only. Not For Use In Diagnostic Procedures.

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
SignalStain is a trademark of Cell Signaling Technology, Inc.

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