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 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 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.

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:

Flow Cytometry (1 Review)
Western Blot (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: 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

Product Datasheets

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COA

SDS

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Product Specific Notices for Human ICAM-1/CD54 Antibody

For research use only

Citations
Reviews

Human ICAM-1/CD54 Antibody

R&D Systems, part of Bio-Techne | Catalog # AF720

Key Product Details

Validated by

Species Reactivity

Validated:

Cited:

Applications