Have any of your S100A9 antibodies been tested for use in neutralization, or blocking assays, on human samples?

Unfortunately at this time we have not tested, or received customer feedback on our S100A9 antibodies for use in blocking or neutralizing assays. If you were planning on testing any of our S100A9 antibodies for neutralizing or blocking capabilities, we would recommend our Innovator’s Reward Program. Under the terms of this program we would be happy to provide a credit for a free vial in exchange for new data on this previously untested or reported application. Please submit this data in the form of an online review. Additional Innovators Reward Program information can be found using this link.

We would like to stain paraffin embedded human intestinal tissue for Calprotectin. One question will be which cells express the majority of Calprotectin - invading neutrophils vs epithelium for example. The antibody list you provide is quite extensive. Which antibody would you recommend for our purpose? Which tissue would you recommend as positive control.

Nine of our antibodies to Calprotectin have been validated for IHC-P with human tissue, and you can see all of these products. A number of our IHC images were generated using human spleen samples, and as such this may be a good choice of a positive control tissue. We do sell a human spleen slide product, which is suitable for IHC-P. Unfortunately I do not have an in-depth knowledge of Calprotectin, however the image shown for our antibody with catalogue number NBP1-02826 demonstrates clear staining of neutrophils. The following paper also suggests that the protein is abundant in neutrophils: PMID 11435495.

Human S100A9 Antibody

Name: Human S100A9 Antibody
Brand: R&D Systems
SKU: AF5578
Rating: 4 (1 reviews)

R&D Systems, part of Bio-Techne | Catalog # AF5578

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Product Specifications
Scientific Data
Applications
Preparation & Storage
Background
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Key Product Details

Validated by

Knockout/Knockdown

Species Reactivity

Human

Applications

Immunocytochemistry, Immunohistochemistry, Intracellular Staining by Flow Cytometry, Knockout Validated, Western Blot

Label

Unconjugated

Antibody Source

Polyclonal Sheep IgG

View all S100A9 Antibodies »

Product Specifications

Immunogen

E. coli-derived recombinant human S100A9
The2-Pro114
Accession # P06702

Specificity

Detects human S100A9 in direct ELISAs and Western blots. In direct ELISAs, less than 5% cross-reactivity with recombinant mouse S100A9 is observed.

Clonality

Polyclonal

Host

Sheep

Isotype

IgG

Scientific Data Images for Human S100A9 Antibody

Detection of Human S100A9 by Western Blot.

Western blot shows lysates of human peripheral blood mononuclear cells (PBMC), human spleen tissue, human tonsil tissue, and human cartilage tissue. PVDF membrane was probed with 0.2 µg/mL of Sheep Anti-Human S100A9 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF5578) followed by HRP-conjugated Anti-Sheep IgG Secondary Antibody (Catalog # HAF016). A specific band was detected for S100A9 at approximately 14 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

S100A9 in MDA‑MB‑468 Human Cell Line.

S100A9 was detected in immersion fixed MDA-MB-468 human breast cancer cell line using Sheep Anti-Human S100A9 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF5578) at 1.7 µg/mL for 3 hours at room temperature. Cells were stained using the NorthernLights™ 557-conjugated Anti-Sheep IgG Secondary Antibody (red; Catalog # NL010) and counterstained with DAPI (blue). Specific staining was localized to cytoplasm. View our protocol for Fluorescent ICC Staining of Cells on Coverslips.

S100A9 in Human Cartilage.

S100A9 was detected in immersion fixed paraffin-embedded sections of human cartilage using Sheep Anti-Human S100A9 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF5578) at 1 µg/mL overnight at 4 °C. Tissue was stained using the Anti-Sheep HRP-DAB Cell & Tissue Staining Kit (brown; Catalog # CTS019) and counterstained with hematoxylin (blue). Specific staining was localized to cytoplasm of chondrocytes. View our protocol for Chromogenic IHC Staining of Paraffin-embedded Tissue Sections.

Western Blot Shows Human S100A9 Specificity by Using Knockout Cell Line.

Western blot shows lysates of MDA-MB-468 human breast cancer parental cell line and S100A9 knock out MDA-MB-468 cell line (KO). PVDF membrane was probed with 0.2 µg/mL of Sheep Anti-Human S100A9 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF5578) followed by HRP-conjugated Anti-Sheep IgG Secondary Antibody (Catalog # HAF016). A specific band was detected for S100A9 at approximately 14 kDa (as indicated) in the parental MDA-MB-468 cell line, but is not detectable in knockout MDA-MB-468 cell line. This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

Detection of S100A9 in Human PBMCs by Flow Cytometry.

Human peripheral blood monocytes (PBMC) were stained with Mouse Anti-Human CD14 PE-conjugated Monoclonal Antibody (Catalog # FAB3832P) and either (A) Sheep Anti-Human S100A9 Polyclonal Antibody (Catalog # AF5578) or (B) Sheep IgG Isotype Control (Catalog # 5-001-A) followed by anti-Sheep IgG APC-conjugated Secondary Antibody (Catalog # F0127). To facilitate intracellular staining, cells were fixed with Flow Cytometry Fixation Buffer (Catalog # FC004) and permeabilized with Flow Cytometry Permeabilization/Wash Buffer (Catalog # FC005). View our protocol for Staining Membrane-associated Proteins.

S100A9 Specificity is Shown by Immunocytochemistry in Knockout Cell Line.

S100A9 was detected in immersion fixed MDA-MB-468 human breast cancer cell line but is not detected in S100A9 knockout (KO) MDA-MB-468 cell line using Sheep Anti-Human S100A9 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF5578) at 1.7 µg/mL for 3 hours at room temperature. Cells were stained using the NorthernLights™ 557-conjugated Anti-Sheep IgG Secondary Antibody (red; Catalog # NL010) and counterstained with DAPI (blue). Specific staining was localized to cytoplasm. View our protocol for Fluorescent ICC Staining of Cells on Coverslips.

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Applications for Human S100A9 Antibody

Application

Recommended Usage

Immunocytochemistry

1-25 µg/mL
Sample: Immersion fixed MDA‑MB‑468 human breast cancer cell line

Immunohistochemistry

5-15 µg/mL
Sample: Immersion fixed paraffin-embedded sections of human cartilage

Intracellular Staining by Flow Cytometry

0.25 µg/10⁶ cells
Sample: Human peripheral blood monocytes (PBMC) fixed with Flow Cytometry Fixation Buffer (Catalog # FC004) and permeabilized with Flow Cytometry Permeabilization/Wash Buffer (Catalog # FC005)

Knockout Validated

S100A9 is specifically detected in MDA‑MB‑468 human breast cancer parental cell line but is not detectable in S100A9 knockout MDA‑MB‑468 cell line.

Western Blot

0.2 µg/mL
Sample: Human peripheral blood mononuclear cells (PBMC), human spleen tissue, human tonsil tissue, and human cartilage tissue

Please Note: Optimal dilutions of this antibody should be experimentally determined.

Reviewed Applications

Read 1 review rated 4 using AF5578 in the following applications:

ELISA (1 Review)

Formulation, Preparation, and Storage

Purification

Antigen Affinity-purified

Reconstitution

Reconstitute at 0.2 mg/mL in sterile PBS. For liquid material, refer to CoA for concentration.

Formulation

Lyophilized from a 0.2 μm filtered solution in PBS with Trehalose. *Small pack size (SP) is supplied either lyophilized or as a 0.2 µm filtered solution in PBS.

Shipping

Lyophilized product is shipped at ambient temperature. Liquid small pack size (-SP) is shipped with polar packs. Upon receipt, store immediately at the temperature recommended below.

Stability & Storage

Use a manual defrost freezer and avoid repeated freeze-thaw cycles.

12 months from date of receipt, -20 to -70 °C as supplied.
1 month, 2 to 8 °C under sterile conditions after reconstitution.
6 months, -20 to -70 °C under sterile conditions after reconstitution.

Background: S100A9

Human S100A9 (also MRP-14, Calgranulin-B, and p14) is a 14 kDa member of the S100 family of EF-hand calcium-binding proteins. It is 114 amino acids (aa) in length and contains short sequential modules. There is an N-terminal helical region, followed by a calcium-binding EF-hand domain, two more helical regions, a second EF-hand domain, and three additional helical regions. S100A9 will noncovalently heterodimerize with S100A8. In the presence of calcium, this heterodimer will form a heterotetramer. S100A9 is expressed in granulocytes, monocytes, and macrophages during acute and chronic inflammation. Human S100A9 shares 62% and 57% aa identity with rat and mouse S100A9, respectively.

Long Name

S100 Calcium Binding Protein A9

Alternate Names

Calgranulin B, MRP-14

Entrez Gene IDs

6280 (Human); 20202 (Mouse)

Gene Symbol

S100A9

UniProt

P06702

Additional S100A9 Products

Product Documents for Human S100A9 Antibody

Product Datasheets

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COA

SDS

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Product Specific Notices for Human S100A9 Antibody

For research use only

Citations
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Protocols
FAQs

Calculators
Flow Cytometry