Human ICAM-1/CD54 Antibody

R&D Systems | Catalog # AF720

R&D Systems

Key Product Details

Validated by

Knockout/Knockdown, Biological Validation

Species Reactivity

Validated:

Human

Cited:

Human

Applications

Validated:

Knockout Validated, Western Blot, Adhesion Blockade, Immunocytochemistry

Cited:

Western Blot, Neutralization, Immunocytochemistry, Bioassay

Label

Unconjugated

Antibody Source

Polyclonal Sheep IgG
View all ICAM-1/CD54 Primary Antibodies »

Product Specifications

Immunogen

Chinese hamster ovary cell line CHO-derived recombinant human ICAM-1
Extracellular domain

Specificity

Selected for its ability to block the adhesion of HSB‑2 cells to immobilized recombinant human (rh) ICAM‑1. In direct ELISAs, approximately 5% cross-reactivity with rhICAM-3 is observed and less than 2% cross-reactivity with rhICAM-2, rhICAM-4, rhICAM-5, recombinant rat ICAM‑1, recombinant mouse ICAM‑1, and rhVCAM-1 is observed.

Clonality

Polyclonal

Host

Sheep

Isotype

IgG

Endotoxin Level

<0.20 EU per 1 μg of the antibody by the LAL method.

Scientific Data Images for Human ICAM-1/CD54 Antibody

Detection of Human ICAM-1/CD54 antibody by Western Blot.

Detection of Human ICAM‑1/CD54 by Western Blot.

Western blot shows lysates of HUVEC human umbilical vein endothelial cells and Ramos human Burkitt's lymphoma cell line untreated (-) or treated (+) with 10 ng/mL Recombinant Human TNF-a (Catalog # 210-TA) for 24 hours. PVDF membrane was probed with 0.2 µg/mL of Sheep Anti-Human ICAM-1/CD54 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF720) followed by HRP-conjugated Anti-Sheep IgG Secondary Antibody (Catalog # HAF016). A specific band was detected for ICAM-1/CD54 at approximately 90 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.
ICAM-1/CD54 antibody in A431 Human Cell Line by Immunocytochemistry (ICC).

ICAM‑1/CD54 in A431 Human Cell Line.

ICAM-1/CD54 was detected in immersion fixed A431 human epithelial carcinoma cell line (left panel; positive stain) and A549 human lung carcinoma cell line (right panel; negative stain) using Sheep Anti-Human ICAM-1/CD54 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF720) 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 plasma membrane. View our protocol for Fluorescent ICC Staining of Cells on Coverslips.

Western Blot Shows Human ICAM‑1/CD54 Specificity by Using Knockout Cell Line.

Western blot shows lysates of Ramos human Burkitt's lymphoma cell line and human ICAM-1/CD54 Ramos human Burkitt's lymphoma cell line (KO). PVDF membrane was probed with 0.2 µg/mL of Sheep Anti-Human ICAM‑1/CD54 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF720) followed by HRP-conjugated Anti-Sheep IgG Secondary Antibody (HAF016). A specific band was detected for ICAM‑1/CD54 at approximately 90 kDa (as indicated) in the parental Ramos human Burkitt's lymphoma cell line, but is not detectable in knockout Ramos human Burkitt's lymphoma cell line. GAPDH (Catalog # AF5718) is shown as a loading control. This experiment was conducted under reducing conditions and using Western Blot Buffer Group 1.
Human ICAM-1 / CD54 ELISA Standard Curve

Human ICAM-1 / CD54 ELISA Standard Curve

Recombinant Human ICAM‑1/CD54 (Catalog # ADP4) was serially diluted and captured by Mouse Anti-Human ICAM‑1/CD54 Monoclonal Antibody (Catalog # MAB720) coated on a Clear Polystyrene Microplate (Catalog # DY990). Sheep Anti-Human ICAM‑1/CD54 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF720) was biotinylated and incubated with the protein captured on the plate. Detection of the standard curve was achieved by incubating Streptavidin-HRP (Catalog # DY998)
Detection of ICAM-1/CD54 by Western Blot

Detection of ICAM-1/CD54 by Western Blot

Immunoblots showing siRNA-mediated knockdown efficiencies corresponding to data of Figure 1.(A) Immunoblots showing knockdown efficiency of siRNA transfections related to Figure 1B. (B) Immunoblots showing knockdown efficiency of siRNA transfections related to Figure 1D. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/30801249), licensed under a CC-BY license. Not internally tested by R&D Systems.

Applications for Human ICAM-1/CD54 Antibody

Application
Recommended Usage

Adhesion Blockade

The adhesion of HSB2 human peripheral blood acute lymphoblastic leukemia cells (5 x 104 cells/well) to immobilized recombinant human ICAM-1/Fc Chimera (12.5 μg/mL, 100 µL/well) was maximally inhibited (80‑100%) by 25 μg/mL of the antibody.

Immunocytochemistry

1-15 µg/mL
Sample: Immersion fixed A431 human epithelial carcinoma cell line

Knockout Validated

ICAM‑1/CD54 is specifically detected in the parental Ramos human Burkitt's lymphoma cell line but is not detectable in knockout Ramos human Burkitt's lymphoma cell line.

Western Blot

0.2 µg/mL
Sample: HUVEC human umbilical vein endothelial cells treated with Recombinant Human TNF‑ alpha (Catalog # 210-TA) and Ramos human Burkitt's lymphoma cell line

Reviewed Applications

Read 2 reviews rated 4.5 using AF720 in the following applications:

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.

Calculators

The reconstitution calculator allows you to quickly calculate the volume of a reagent to reconstitute your vial. Simply enter the mass of reagent and the target concentration and the calculator will determine the rest.

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Background: ICAM-1/CD54

Intercellular Adhesion Molecule-1 (ICAM-1) binds the leukocyte integrins LFA-1 and Mac-1. ICAM-1 expression is weak on leukocytes, epithelial and resting endothelial cells, as well as some other cell types, but expression can be stimulated by IFN-gamma, TNF-alpha, IL-1 beta and LPS. Soluble ICAM-1 is found in a biologically active form in serum, probably as a result of proteolytic cleavage from the cell surface, and is elevated in patients with various inflammatory syndromes such as septic shock, LAD, cancer and transplantation.

Long Name

Intercellular Adhesion Molecule 1

Alternate Names

CD54, ICAM1

Entrez Gene IDs

3383 (Human); 15894 (Mouse); 25464 (Rat)

Gene Symbol

ICAM1

Additional ICAM-1/CD54 Products

Product Documents for Human ICAM-1/CD54 Antibody

Certificate of Analysis

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Note: Certificate of Analysis not available for kit components.

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Product Specific Notices for Human ICAM-1/CD54 Antibody

For research use only

Citations for Human ICAM-1/CD54 Antibody

Customer Reviews for Human ICAM-1/CD54 Antibody (2)

4.5 out of 5
2 Customer Ratings
5 Stars
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4 Stars
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  • Name: Anonymous
    Application: Western Blot
    Sample Tested: Purified protein
    Species: Mouse
    Verified Customer | Posted 03/23/2018
  • Name: Anonymous
    Application: Flow Cytometry
    Sample Tested: See PMID 20022380
    Species: Human
    Verified Customer | Posted 01/09/2015

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