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66297
Arginase-1 (D4E3M™) XP® Rabbit mAb (Alexa Fluor® 488 Conjugate)
Antibody Conjugates
Monoclonal Antibody
R
Recombinant

Arginase-1 (D4E3M) XP® Rabbit mAb (Alexa Fluor® 488 Conjugate) #66297

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  1. IF
Immunofluorescence Image 1: Arginase-1 (D4E3M™) XP® Rabbit mAb (Alexa Fluor® 488 Conjugate)
Confocal immunofluorescent analysis of fixed frozen mouse liver (left) and mouse pancreas (right), labeled with Arginase-1 (D4E3M) XP® Rabbit mAb (Alexa Fluor® 488 Conjugate) #66297 (green) and DAPI #4083 (blue).
Immunofluorescence Image 2: Arginase-1 (D4E3M™) XP® Rabbit mAb (Alexa Fluor® 488 Conjugate)
Confocal immunofluorescent analysis of fixed frozen mouse liver (left) and mouse pancreas (right), labeled with Arginase-1 (D4E3M) XP® Rabbit mAb (Alexa Fluor® 488 Conjugate) #66297 (green) and DAPI #4083 (blue).
Immunofluorescence Image 1: Arginase-1 (D4E3M™) XP® Rabbit mAb (Alexa Fluor® 488 Conjugate)
Confocal immunofluorescent analysis of mouse primary bone marrow-derived macrophages (BMDMs) using Arginase-1 (D4E3M) XP® (Alexa Fluor® 488 Conjugate) Rabbit mAb (green). BMDMs were differentiated with M-CSF (20 ng/ml, 7 days) and activated with either IL-4/cAMP (20 ng/ml, 0.5 mM, 24 hours; left) or LPS/IFNγ (50 ng/ml, 20 ng/ml, 24 hours; right). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).
To Purchase # 66297
製品# サイズ 数量 価格 在庫
66297S
100 µl  (50 tests)

Supporting Data

REACTIVITY H M R
SENSITIVITY Endogenous
MW (kDa)
Source/Isotype Rabbit IgG

Application Key:

  • WB-Western Blot
  • IP-Immunoprecipitation
  • IHC-Immunohistochemistry
  • ChIP-Chromatin Immunoprecipitation
  • C&R-CUT&RUN
  • C&T-CUT&Tag
  • DB-Dot Blot
  • eCLIP-eCLIP
  • IF-Immunofluorescence
  • F-Flow Cytometry

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
  • GP-Guinea Pig
  • Rab-Rabbit
  • All-All Species Expected

Product Description

This Cell Signaling Technology antibody is conjugated to Alexa Fluor® 488 fluorescent dye and tested in-house for direct flow cytometric analysis in human cells. This antibody is expected to exhibit the same species cross-reactivity as the unconjugated Arginase-1 (D4E3M™) XP® Rabbit mAb #93668.

Product Usage Information

Application Dilution
Immunofluorescence (Frozen) 1:100
Immunofluorescence (Immunocytochemistry) 1:100 - 1:200

Storage

Supplied in PBS (pH 7.2), less than 0.1% sodium azide and 2 mg/ml BSA. Store at 4°C. Do not aliquot the antibody. Protect from light. Do not freeze.

Protocol

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Immunofluorescence (Frozen)

A. Solutions and Reagents

NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalently purified water.

  1. 20X Phosphate Buffered Saline (PBS): (9808) To prepare 1L 1X PBS: add 50 ml 20X PBS to 950 ml dH2O, mix. Adjust pH to 8.0.
  2. Formaldehyde: 16%, methanol free, Polysciences, Inc. (cat# 18814), use fresh and store opened vials at 4°C in dark, dilute in 1X PBS for use.
  3. Methanol, 100%
  4. Blocking Buffer (1X PBS / 5% normal goat serum (#5425) / 0.3% Triton™ X-100): To prepare 10 ml: add 0.5 ml normal goat serum and 0.5 ml 20X PBS to 9.0 ml dH2O, mix. While stirring, add 30 µl Triton™ X-100.
  5. Antibody Dilution Buffer (1X PBS / 1% BSA / 0.3% Triton X-100): To prepare 10 ml, add 30 µl Triton™ X-100 to 10 ml 1X PBS. Mix well then add 0.1 g BSA (9998), mix.
  6. Prolong® Gold AntiFade Reagent (#9071), Prolong® Gold AntiFade Reagent with DAPI (#8961).

B. Specimen Preparation - Frozen/Cryostat Sections (IF-F)

  1. For fixed frozen tissue proceed with Immunostaining (Section C).
  2. For fresh, unfixed frozen tissue, please fix immediately, as follows:
    1. Cover sections with 4% formaldehyde dilute in 1X PBS.

      NOTE: Formaldehyde is toxic, use only in fume hood.

    2. Allow sections to fix for 15 minutes at room temperature.
    3. Aspirate liquid, rinse three times in 1X PBS for 5 minutes each.
    4. Proceed with Immunostaining (Section C).

C. Immunostaining

NOTE: All subsequent incubations should be carried out at room temperature unless otherwise noted in a humid light-tight box or covered dish/plate to prevent drying and fluorochrome fading.

  1. Methanol Permeabilization Step: Cover tissue sections with ice-cold 100% methanol (use enough to cover completely to a depth of 3–5 mm, DO NOT LET DRY), incubate in methanol for 10 minutes at –20°C, rinse in 1X PBS for 5 minutes.
  2. Block specimen in Blocking Buffer for 60 minutes.
  3. While blocking, prepare primary antibody by diluting as indicated on protocol on product webpage in Antibody Dilution Buffer.
  4. Aspirate blocking solution, apply diluted primary antibody.
  5. Incubate overnight at 4°C.
  6. Rinse three times in 1X PBS for 5 minutes each.
  7. Coverslip slides with Prolong® Gold Antifade Reagent (#9071), Prolong® Gold AntiFade Reagent with DAPI (#8961).
  8. For best results examine specimens immediately using appropriate excitation wavelength. For long term storage, store slides flat at 4°C protected from light.

posted November 2006

revised December 2010

Protocol Id: 464

Immunofluorescence (Immunocytochemistry)

A. Solutions and Reagents

NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalently purified water.

  1. 20X Phosphate Buffered Saline (PBS): (9808) To prepare 1L 1X PBS: add 50 ml 20X PBS to 950 ml dH2O, mix. Adjust pH to 8.0.
  2. Formaldehyde: 16%, methanol free, Polysciences, Inc. (cat# 18814), use fresh and store opened vials at 4°C in dark, dilute in 1X PBS for use.
  3. Methanol, 100%
  4. Blocking Buffer (1X PBS / 5% normal goat serum (#5425) / 0.3% Triton™ X-100): To prepare 10 ml: add 0.5 ml normal goat serum and 0.5 ml 20X PBS to 9.0 ml dH2O, mix. While stirring, add 30 µl Triton™ X-100.
  5. Antibody Dilution Buffer (1X PBS / 1% BSA / 0.3% Triton X-100): To prepare 10 ml, add 30 µl Triton™ X-100 to 10 ml 1X PBS. Mix well then add 0.1 g BSA (9998), mix.
  6. Prolong® Gold AntiFade Reagent (#9071), Prolong® Gold AntiFade Reagent with DAPI (#8961).

B. Specimen Preparation - Cultured Cell Lines (IF-IC)

NOTE: Cells should be grown, treated, fixed and stained directly in multiwell plates, chamber slides or on coverslips.

  1. Aspirate liquid, then cover cells to a depth of 2–3 mm with 4% formaldehyde in 1X PBS.
    NOTE: Formaldehyde is toxic, use only in fume hood.
  2. Allow cells to fix for 15 minutes at room temperature.
  3. Aspirate fixative, rinse three times in 1X PBS for 5 minutes each.
  4. Proceed with Immunostaining (Section C).

C. Immunostaining

NOTE: All subsequent incubations should be carried out at room temperature unless otherwise noted in a humid light-tight box or covered dish/plate to prevent drying and fluorochrome fading.

  1. Methanol Permeabilization Step: Cover cells with ice-cold 100% methanol (use enough to cover completely to a depth of 3–5 mm, DO NOT LET DRY), incubate in methanol for 10 minutes at –20°C, rinse in 1X PBS for 5 minutes.
  2. Block specimen in Blocking Buffer for 60 minutes.
  3. While blocking, prepare primary antibody by diluting as indicated on product webpage in Antibody Dilution Buffer.
  4. Aspirate blocking solution, apply diluted primary antibody.
  5. Incubate overnight at 4°C.
  6. Rinse three times in 1X PBS for 5 minutes each.
  7. Coverslip slides with Prolong® Gold Antifade Reagent (#9071), Prolong® Gold AntiFade Reagent with DAPI (#8961).
  8. For best results examine specimens immediately using appropriate excitation wavelength. For long term storage, store slides flat at 4°C protected from light.

posted November 2006

revised December 2010

Protocol Id: 220

Specificity / Sensitivity

Arginase-1 (D4E3M™) XP® Rabbit mAb (Alexa Fluor® 488 Conjugate) recognizes endogenous levels of total arginase-1 protein. This antibody does not cross-react with arginase-2.

Species Reactivity:

Human, Mouse, Rat

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Val47 of human arginase-1 protein.

Background

L-arginine plays a critical role in regulating the immune system (1-3). In inflammation, cancer, and certain other pathological conditions, myeloid cell differentiation is inhibited leading to a heterogeneous population of immature myeloid cells, known as myeloid-derived suppressor cells (MDSCs). MDSCs are recruited to sites of cancer-associated inflammation and express high levels of arginase-1 (4). Arginase-1 catalyzes the final step of the urea cycle converting L-arginine to L-ornithine and urea (5). Thus, MDSCs increase the catabolism of L-arginine resulting in L-arginine depletion in the inflammatory microenvironment of cancer (4,6). The reduced availability of L-arginine suppresses T cell proliferation and function and thus contributes to tumor progression (4,6). Arginase-1 is of great interest to researchers looking for a therapeutic target to inhibit the function of MDSCs in the context of cancer immunotherapy (7). In addition, research studies have demonstrated that arginase-1 distinguishes primary hepatocellular carcinoma (HCC) from metastatic tumors in the liver, indicating its value as a potential biomarker in the diagnosis of HCC (8,9).

使用に関する制限

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For Research Use Only. Not for Use in Diagnostic Procedures.
Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
Alexa Fluor is a registered trademark of Life Technologies Corporation.
This product is provided under an intellectual property license from Life Technologies Corporation. The transfer of this product is conditioned on the buyer using the purchased product solely in research conducted by the buyer, excluding contract research or any fee for service research, and the buyer must not (1) use this product or its components for (a) diagnostic, therapeutic or prophylactic purposes; (b) testing, analysis or screening services, or information in return for compensation on a per-test basis; or (c) manufacturing or quality assurance or quality control, and/or (2) sell or transfer this product or its components for resale, whether or not resold for use in research. For information on purchasing a license to this product for purposes other than as described above, contact Life Technologies Corporation, 5791 Van Allen Way, Carlsbad, CA 92008 USA or [email protected]
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