IL-17A Antibody, anti-human, REAfinity™

Name: IL-17A Antibody, anti-human, REAfinity™
Availability: InStock

IL-17A Antibody, anti-human, REAfinity™Extended ValidationRecombinant

Clone:

REA1063

Type of antibody:

Primary antibodies, Recombinant antibodies

Isotype:

recombinant human IgG1

Applications:

ICFC

Alternative names:

CTLA-8, IL-17

Intracellular flow cytometry applications forIL-17A Antibody, anti-human, REAfinity™

APC
APC-Vio 770
FITC
PE
PE-Vio 615
PE-Vio 770
Vio B515

Conjugate:

APC

Recommended dilution:

1:50

Remark:

The antibody is suited for staining of formaldehyde-fixed cells.

Tested:

QC tested

Products used:

130-118-398, 130-118-244

Protocols:

Intracellular flow cytometry staining protocol (Inside Stain Kit 1:50)

Description:

Naive CD4⁺ T cells were isolated from human peripheral blood mononuclear cells (PBMCs). Cells were either left unstimulated (left images) or stimulated with PMA (50 ng/mL) and Ionomycin (1 µg/mL) for six hours. After two hours, brefeldin A (1 µg/mL) was added to the stimulated and to the unstimulated cells. Cells were fixed, permeabilized, and intracellularly stained with Anti-IL-17A antibodies as well as with CD3 antibodies. Cells were then analyzed by flow cytometry using the MACSQuant^® Analyzer. The Tandem Signal Enhancer has been used to increase binding specificity of tandem-dye–conjugated antibodies. Cell debris were excluded from the analysis based on scatter signals.

Extended validation for IL-17A Antibody, anti-human, REAfinity™

Specificity

Epitope competition

In order to compare the epitope specificity of an antibody, the clone being used is compared with other known clones recognizing the same antigen in a competition assay.

Other clones	Overlap in epitope recognition with REA1063
CZ8-23G1	++
BL168	++
N49-653	++
eBio64DEC17	-
Cells were incubated with an excess of purified unconjugated IL-17A (REA1063) antibody followed by staining with fluorochrome-conjugated antibodies of other known clones against the same marker. Based on the fluorescence signal obtained, the clones were identified as recognizing completely overlapping (++), partially overlapping (+), or completely different epitopes (-) of the marker.

Sensitivity

Performance comparison

Selected fluorochrome conjugated antibodies from Miltenyi Biotec were compared to commercially available hybridoma clones in flow cytometry analysis.

Flow cytometric comparison of different clones for IL-17A. Human peripheral blood mononuclear cells (PBMCs) were stimulated with 20 ng/mL Phorbol 12-myristate 13-acetate (PMA) and 1 µg/ml Ionomycin for 1 hour, followed by an incubation with 1µg/mL Brefeldin A for 4 hours. Cells were stained with a suitable counterstaining and then fixed and permeabilized followed by a staining with IL-17A antibodies. As a control, IL-17A antibody staining was omitted and cells were measured in the same channels. Flow cytometry was performed with the MACSQuant^® Analyzer. CD3⁺/CD4⁺ cells were pregated for the analysis. Cell debris and cell doublets were excluded from the analysis based on scatter signals. No FcR Blocking Reagent was used. The recommended titers of respective antibodies from different suppliers were used.

Flow cytometric comparison of different clones for IL-17A. Human peripheral blood mononuclear cells (PBMCs) were stimulated with 20 ng/mL Phorbol 12-myristate 13-acetate (PMA) and 1 µg/ml Ionomycin for 1 hour, followed by an incubation with 1µg/mL Brefeldin A for 4 hours. Cells were stained with a suitable counterstaining and then fixed and permeabilized followed by a staining with IL-17A antibodies. As a control, IL-17A antibody staining was omitted and cells were measured in the same channels. Flow cytometry was performed with the MACSQuant^® Analyzer. CD3⁺/CD4⁺ cells were pregated for the analysis. Cell debris and cell doublets were excluded from the analysis based on scatter signals. No FcR Blocking Reagent was used. The recommended titers of respective antibodies from different suppliers were used.

Flow cytometric comparison of different clones for IL-17A. Human peripheral blood mononuclear cells (PBMCs) were stimulated with 20 ng/mL Phorbol 12-myristate 13-acetate (PMA) and 1 µg/ml Ionomycin for 1 hour, followed by an incubation with 1µg/mL Brefeldin A for 4 hours. Cells were stained with a suitable counterstaining and then fixed and permeabilized followed by a staining with IL-17A antibodies. As a control, IL-17A antibody staining was omitted and cells were measured in the same channels. Flow cytometry was performed with the MACSQuant^® Analyzer. CD3⁺/CD4⁺ cells were pregated for the analysis. Cell debris and cell doublets were excluded from the analysis based on scatter signals. No FcR Blocking Reagent was used. The recommended titers of respective antibodies from different suppliers were used.

Flow cytometric comparison of different clones for IL-17A. Human peripheral blood mononuclear cells (PBMCs) were stimulated with 20 ng/mL Phorbol 12-myristate 13-acetate (PMA) and 1 µg/ml Ionomycin for 1 hour, followed by an incubation with 1µg/mL Brefeldin A for 4 hours. Cells were stained with a suitable counterstaining and then fixed and permeabilized followed by a staining with IL-17A antibodies. As a control, IL-17A antibody staining was omitted and cells were measured in the same channels. Flow cytometry was performed with the MACSQuant^® Analyzer. CD3⁺/CD4⁺ cells were pregated for the analysis. Cell debris and cell doublets were excluded from the analysis based on scatter signals. No FcR Blocking Reagent was used. The recommended titers of respective antibodies from different suppliers were used.

Flow cytometric comparison of different clones for IL-17A. Human peripheral blood mononuclear cells (PBMCs) were stimulated with 20 ng/mL Phorbol 12-myristate 13-acetate (PMA) and 1 µg/ml Ionomycin for 1 hour, followed by an incubation with 1µg/mL Brefeldin A for 4 hours. Cells were stained with a suitable counterstaining and then fixed and permeabilized followed by a staining with IL-17A antibodies. As a control, IL-17A antibody staining was omitted and cells were measured in the same channels. Flow cytometry was performed with the MACSQuant^® Analyzer. CD3⁺/CD4⁺ cells were pregated for the analysis. Cell debris and cell doublets were excluded from the analysis based on scatter signals. No FcR Blocking Reagent was used. The recommended titers of respective antibodies from different suppliers were used.

Flow cytometric comparison of different clones for IL-17A. Human peripheral blood mononuclear cells (PBMCs) were stimulated with 20 ng/mL Phorbol 12-myristate 13-acetate (PMA) and 1 µg/ml Ionomycin for 1 hour, followed by an incubation with 1µg/mL Brefeldin A for 4 hours. Cells were stained with a suitable counterstaining and then fixed and permeabilized followed by a staining with IL-17A antibodies. As a control, IL-17A antibody staining was omitted and cells were measured in the same channels. Flow cytometry was performed with the MACSQuant^® Analyzer. CD3⁺/CD4⁺ cells were pregated for the analysis. Cell debris and cell doublets were excluded from the analysis based on scatter signals. No FcR Blocking Reagent was used. The recommended titers of respective antibodies from different suppliers were used.

Specifications for IL-17A Antibody, anti-human, REAfinity™

Overview

Clone REA1063 recognizes human interleukin 17A (IL-17A). IL-17A, also known as CTLA8, is a member of the IL-17 family (IL-17A–F), is a disulfide-linked homodimeric glycoprotein. IL-17A is produced by CD4

⁺

T helper (Tʜ) cells, a third T cell subset termed Tʜ17, which secrete also cytokines such as IL-17F and IL-22 and express the natural killer (NK) cell marker CD161. IL-17A secretion has also been described for other cell types, such as CD8

⁺

memory T cells. Furthermore, intracellular IL-17A has also been detected in eosinophils, neutrophils, and blood monocytes. Emerging data about Tʜ17 cells suggest that these cells are involved in the recruitment of neutrophils to control early stages of infections to a number of pathogens, such as extracellular bacteria and fungi. IL-17A and Tʜ17 cells have been shown to play an important role in immune-mediated diseases, such as rheumatoid arthritis, psoriasis, multiple sclerosis, asthma, inflammatory bowel diseases, and other immune-mediated inflammatory conditions.

Depending on the cytokine milieu present at time of the initial engagement, CD4

⁺

naive T cells can differentiate into various subsets (Tʜ1, Tʜ2, and Tʜ17). For the differentiation into Tʜ17 cells several cytokines have been described, including TGF-β, IL-1β, IL-6, IL-21, and IL-23. RORγt was identified as the master regulator gene for Tʜ17 cells.

Additional information: Clone REA1063 displays negligible binding to Fc receptors.

Alternative names

CTLA-8, IL-17

Detailed product information

Technical specifications

Clone	REA1063
Clonality	monoclonal
Isotype	recombinant human IgG1
Isotype control	REA Control Antibody (I), human IgG1
Host	human cell line
Type of antibody	Primary antibodies, Recombinant antibodies
Species	human
Antigen	IL-17A
Alternative names of antigen	CTLA-8, IL-17
Molecular mass of antigen [kDa]	15
Distribution of antigen	monocytes, T cells, CD4+ T cells, CD8+ T cells, eosinophils, neutrophils, Th17 cells
Entrez Gene ID	3605
RRID	AB_2751465, AB_2751500, AB_2751466, AB_2751501, AB_2751467, AB_2751502, AB_2751468, AB_2751503, AB_2751469, AB_2751504, AB_2751470, AB_2751505, AB_2751471, AB_2751499

Resources for IL-17A Antibody, anti-human, REAfinity™

Certificates

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to search for Certificates of Analysis (CoA) by lot number.

References for IL-17A Antibody, anti-human, REAfinity™

Publications

Yao, Z. et al. (1995) Human IL-17: a novel cytokine derived from T cells. J Immunol 155: 5483-5486
Kolls, J. K. and Linden, A. (2004) Interleukin-17 family members and inflammation. Immunity 21: 467-476
Ivanov, I. I. et al. (2006)
The orphan nuclear receptor RORγt directs the differentiation program of proinflammatory IL-17
⁺
T helper cells.
Cell 126: 1121-1133
Cosmi, L. et al. (2008)
Human interleukin 17–producing cells originate from a CD161
⁺
CD4
⁺
T cell precursor.
J. Exp. Med. 205: 1903-1916
Manel, H. et al. (2008) The differentiation of human Tʜ-17 cells requires transforming growth factor-β and induction of the nuclear receptor RORγt. Nat Immunol 9: 641-649
Tesmer, L. A. et al. (2008) Tʜ17 cells in human disease. Immunol. Rev. 223: 87-113
Volpe, E. et al. (2008) A critical function for transforming growth factor-β, interleukin 23 and proinflammatory cytokines in driving and modulating human Tʜ-17 responses. Nat Immunol 9: 650-657
Yang, L. et al. (2008) IL-21 and TGF-beta are required for differentiation of human Tʜ17 cells. Nature 454: 350-352

Applications

Extended validation