CD25 antibody (AA 201-268)

Catalog No. ABIN670371

This anti-CD25 antibody is a Rabbit Polyclonal antibody detecting CD25 in FACS, WB, ELISA and IHC (p). Suitable for Human and Mouse. Independently validated for use in Immunohistochemistry.

Quick Overview for CD25 antibody (AA 201-268) (ABIN670371)

Target: CD25 (IL2RA) (Interleukin 2 Receptor, alpha (IL2RA))

Reactivity: Human, Mouse

Host: Rabbit

Clonality: Polyclonal

Conjugate: This CD25 antibody is un-conjugated

Application: Flow Cytometry (FACS), Western Blotting (WB), ELISA, Immunohistochemistry (Paraffin-embedded Sections) (IHC (p))

Product Details anti-CD25 Antibody

Binding Specificity: AA 201-268

Cross-Reactivity: Human, Mouse

Predicted Reactivity: Rat

Purification: Purified by Protein A.

Immunogen: KLH conjugated synthetic peptide derived from mouse CD25/IL2RA

Isotype: IgG

Application Details

Application Notes: WB 1:300-5000
ELISA 1:500-1000
FCM 1:20-100
IHC-P 1:200-400

Restrictions: For Research Use only

Independent Validation (IHC)

Validation #104539 (Immunohistochemistry)

by: TRON

No.: #104539

Date: 02/28/2024

Antigen: CD25

Lot Number: AG10271265

Method validated: Immunohistochemistry

Positive Control:

FFPE human tonsil

Notes:

Passed. The CD25 antibody ABIN670371 recognizes CD25 in human tonsil tissue.

Validation Images

IIHC staining of human tonsil with anti-CD25 antibody ABIN670371 (1:250) and counterstain with hematoxylin.

IHC staining of human tonsil with anti-CD25 antibody ABIN670371 (1:500) and counterstain with hematoxylin.

Full Methods

Primary Antibody: ABIN670371

Secondary Antibody: HRP-conjugated goat anti-rabbit antibody (medac diagnostics, DPVR-110HRP, Lot 101122)

Full Protocol:

• Dewax and rehydrate tissue sections on glass slides in a descending xylene and graded alcohol series:
• • xylene 2x for 5 min.
  • 100 % (v/v) ethanol 2x for 5 min.
  • 96 % (v/v) ethanol 2x for 5 min.
  • 70 % (v/v) ethanol 2x for 5 min.
• Wash slides 2x 5 min with ddH₂O.
• Cook slides for 30 min at 95 °C in citrate buffer pH 6.
• Wash slides 2x 3 min with ddH₂O.
• Incubate slides in 0.3 % (v/v) H₂O₂ in PBS for 15 min to quench endogenous peroxidase.
• Wash slides 3x 3 min in 1X PBS.
• Block slides in 10 % goat serum in PBS for 30 min at RT.
• Incubate slides with primary rabbit anti-CD25 antibody (antibodies-online, ABIN670371, lot AG10271265) diluted 1:250 and 1:500 in 10 % goat serum in PBS ON at 4 °C in a humidified chamber.
• Wash slides 3x 3 min in 1X PBS.
• Incubate slides with secondary HRP-conjugated goat anti-rabbit antibody (medac diagnostics, DPVR-110HRP, Lot 101122) ready-to-use in 10 % goat serum in PBS for 30 min at RT in a humidified chamber.
• Wash slides 3x 3 min in 1X PBS.
• HRP development.
• Counterstain slides with hematoxylin solution (Carl Roth, T865.2).
• Wash slides for 15 min under running tap water.
• Dehydrate sections in an ascending alcohol-xylene series:
• • 70% ethanol 2x for 10 sec.
  • 96% ethanol 2x for 10 sec.
  • 100% ethanol 2x for 10 sec.
  • xylene 2x for 10 sec.
• Mount sections with Entellan new mounting medium (Sigma-Aldrich, 1008690500).
• Dry sections.
• Image acquisition on an Aperio CS2 Digital Pathology Slide Scanner (Leica), Magnification 40x.

Experimental Notes:

• ABIN670371 appears to successfully recognize CD25 in human tonsils, but in mouse spleen and lung, the signal is not present where expected.

• ABIN670371 does not appear to recognize CD25 in mouse samples. CD25 should be highly expressed on Tregs, which by definition express FoxP3. ABIN670371 signal does not co-localize with FoxP3 in either spleen or lung. Furthermore, in both tissues, the signal from ABIN670371 is nuclear, whereas it is expected to be membrane. ABIN670371 signal does not co-localize with CD8/CD4 in either spleen or lung although CD25 should (at least partially) co-localize with CD8 and CD4.

Handling

Format: Liquid

Concentration: 1 μg/μL

Buffer: 0.01M TBS( pH 7.4) with 1 % BSA, 0.02 % Proclin300 and 50 % Glycerol.

Preservative: ProClin

Precaution of Use: This product contains ProClin: a POISONOUS AND HAZARDOUS SUBSTANCE, which should be handled by trained staff only.

Storage: 4 °C,-20 °C

Storage Comment: Shipped at 4°C. Store at -20°C for one year. Avoid repeated freeze/thaw cycles.

Expiry Date: 12 months

Target Details for CD25

Target: CD25 (IL2RA) (Interleukin 2 Receptor, alpha (IL2RA))

Alternative Name: IL2RA

Background:

Synonyms: CD25, Il2r, Ly-43, Interleukin-2 receptor subunit alpha, IL-2 receptor subunit alpha, IL-2-RA, IL-2R subunit alpha, IL2-RA, p55, Il2ra

Background: The interleukin 2 (IL2) receptor alpha (IL2RA) and beta (IL2RB) chains, together with the common gamma chain (IL2RG), constitute the high-affinity IL2 receptor. Homodimeric alpha chains (IL2RA) result in low-affinity receptor, while homodimeric beta (IL2RB) chains produce a medium-affinity receptor. Normally an integral-membrane protein, soluble IL2RA has been isolated and determined to result from extracellular proteolyisis. Alternately-spliced IL2RA mRNAs have been isolated, but the significance of each is presently unknown. Mutations in this gene are associated with interleukin 2 receptor alpha deficiency.[provided by RefSeq, Nov 2009].

Gene ID: 16184

UniProt: P01590

Pathways: JAK-STAT Signaling, Growth Factor Binding, Activated T Cell Proliferation