NCAM-1/CD56 Antibody

R&D Systems | Catalog # AF2408

R&D Systems

Key Product Details

Validated by

Knockout/Knockdown

Species Reactivity

Validated:

Human, Mouse

Cited:

Human, Mouse, Xenograft

Applications

Validated:

Knockout Validated, Western Blot, Flow Cytometry, Immunocytochemistry, Simple Western, CyTOF-ready

Cited:

Immunohistochemistry, Immunohistochemistry-Paraffin, Western Blot, Flow Cytometry, Immunocytochemistry, Microarray

Label

Unconjugated

Antibody Source

Polyclonal Goat IgG
View all NCAM-1/CD56 Primary Antibodies »

Product Specifications

Immunogen

Mouse myeloma cell line NS0-derived recombinant human NCAM1/CD56
Leu20-Pro603
Accession # NP_001070150

Specificity

Detects human NCAM-1/CD56 in direct ELISAs and Western blots. In direct ELISAs and Western blots, less than 1% cross‑reactivity with recombinant human (rh) ALCAM, rhBCAM and rhEpCAM is observed.

Clonality

Polyclonal

Host

Goat

Isotype

IgG

Scientific Data Images for NCAM-1/CD56 Antibody

Detection of Human and Mouse NCAM-1/CD56 antibody by Western Blot.

Detection of Human and Mouse NCAM‑1/CD56 by Western Blot.

Western blot shows lysates of human brain (cerebellum and motor cortex) tissue and mouse brain (cerebellum) tissue. PVDF membrane was probed with 0.5 µg/mL of Goat Anti-Human/Mouse NCAM-1/CD56 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF2408) followed by HRP-conjugated Anti-Goat IgG Secondary Antibody (Catalog # HAF017). Specific bands were detected for NCAM-1/CD56 at approximately 100-150 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.
NCAM-1/CD56 antibody in SH-SY5Y Human Neuroblastoma Cells by Immunocytochemistry (ICC).

NCAM-1/CD56 in SH-SY5Y Human Neuroblastoma Cells.

SH-SY5Y human neuroblastoma cells were cultured overnight in the presence of 1 μM Retinoic Acid (0695/50) prior to immersion fixation. Neural Cell Adhesion Molecule 1 (NCAM-1)/CD56 was detected using a Goat Anti-Human/Mouse NCAM-1/CD56 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF2408). The cells were stained with the NorthernLights 557-conjugated Donkey Anti-Goat IgG Affinity-purified Secondary Antibody (red; Catalog # NL001). Actin filaments were stained with FITC-conjugated Phalloidin (green) and cell nuclei were counter-stained with DAPI (blue). NCAM-1/CD56 immuno-reactivity was localized to the plasma membrane. View our protocol for Fluorescent ICC Staining of Cells on Coverslips.
NCAM-1/CD56 antibody in BG01V Human Embryonic Stem Cells by Immunocytochemistry (ICC).

NCAM‑1/CD56 in BG01V Human Embryonic Stem Cells.

NCAM-1/CD56 was detected in immersion fixed BG01V human embryonic stem cells differentiated into neural progenitor cells using Goat Anti-Human/Mouse NCAM-1/CD56 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF2408) at 10 µg/mL for 3 hours at room temperature. Cells were stained using the NorthernLights™ 557-conjugated Anti-Goat IgG Secondary Antibody (red; Catalog # NL001) and counterstained with DAPI (blue). Specific staining was localized to cytoplasm. View our protocol for Fluorescent ICC Staining of Stem Cells on Coverslips.
Detection of Human and Mouse NCAM-1/CD56 antibody by Simple WesternTM.

Detection of Human and Mouse NCAM‑1/CD56 by Simple WesternTM.

Simple Western lane view shows lysates of human and mouse brain (cerebellum) tissue, loaded at 0.2 mg/mL. A specific band was detected for NCAM-1/CD56 at approximately 143 kDa (as indicated) using 5 µg/mL for human lysates and 25 µg/mL for mouse lysates of Goat Anti-Human/Mouse NCAM-1/ CD56 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF2408) followed by 1:50 dilution of HRP-conjugated Anti-Goat IgG Secondary Antibody (Catalog # HAF109). This experiment was conducted under reducing conditions and using the 12-230 kDa separation system.

Western Blot Shows Human NCAM‑1/CD56 Specificity by Using Knockout Cell Line.

Western blot shows lysates of U937 human histiocytic lymphoma cell line and human NCAM-1 knockout U937 human histiocytic lymphoma cell line (KO). PVDF membrane was probed with 0.25 µg/mL of Goat Anti-Human/Mouse NCAM‑1/CD56 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF2408) followed by HRP-conjugated Anti-Goat IgG Secondary Antibody (HAF017). A specific band was detected for NCAM‑1/CD56 at approximately 160 kDa (as indicated) in the parental U937 human histiocytic lymphoma cell line, but is not detectable in knockout U937 human histiocytic lymphoma cell line. GAPDH (AF5718) is shown as a loading control. This experiment was conducted under reducing conditions and using Western Blot Buffer Group 1.
Human NCAM-1 / CD56 ELISA Standard Curve

Human NCAM-1 / CD56 ELISA Standard Curve

Recombinant Human NCAM‑1/CD56 120 isoform (Catalog # 2408-NC) was serially diluted and captured by Mouse Anti-Human/Primate NCAM‑1/CD56 Monoclonal Antibody (Catalog # MAB2408) coated on a Clear Polystyrene Microplate (Catalog # DY990). Goat Anti-Human/Mouse NCAM‑1/CD56 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF2408) was biotinylated and incubated with the protein captured on the plate. Detection of the standard curve was achieved by incubating Streptavidin-HRP (Catalog # DY998)
Human NCAM-1/CD56 DuoSet ELISA

Human NCAM-1/CD56 DuoSet ELISA

Recombinant Human NCAM‑1/CD56 120 isoform (Catalog # 2408-NC) was serially diluted and captured by Mouse Anti-Human/Primate NCAM‑1/CD56 Monoclonal Antibody (Catalog # MAB2408) coated on a Clear Polystyrene Microplate (Catalog # DY990). Goat Anti-Human/Mouse NCAM‑1/CD56 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF2408) 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 NCAM-1/CD56 by Western Blot

Detection of NCAM-1/CD56 by Western Blot

GDE2 overexpression promotes EV release in HEK293T cells. (A) Schematic of the workflow used to produce, purify, and analyze EVs from HEK293T cells, along with representative TEM image of purified EVs (scale bar = 100 nm). (B) Representative Western blots displaying markers, NCAM-1, Alix, and Annexin A1 in cell lysates and EVs from transfected HEK293T cells. (C) Quantification of fold change (FC) for each marker analyzed, normalized to signal derived from corresponding cell lysate. DMEM, Dulbecco’s modified Eagle medium; EV, extracellular vesicle; HEK293T, Human Embryonic Kidney 293T; NCAM-1, neural cell adhesion molecule-1; NT, non-transfected; SEC, size exclusion chromatography; UC, ultracentrifugation; ns p > 0.05, * p < 0.05, ** p < 0.01; *** p < 0.001; mean ± SEM, one-way ANOVA with Tukey’s multiple comparisons test. N = 3 biological replicates. Image collected and cropped by CiteAb from the following open publication (https://www.mdpi.com/2073-4409/13/17/1414), licensed under a CC-BY license. Not internally tested by R&D Systems.
Detection of NCAM-1/CD56 by Flow Cytometry

Detection of NCAM-1/CD56 by Flow Cytometry

Cluster of differentiation (CD)56 expression by human tumor cell lines. (A) Flow cytometric analysis of tumor cells labelled with CD56-PE (black line) or corresponding isotype control (filled grey), represented as histogram overlays. (B) Real-time qPCR data of the expression levels of the different CD56 isoforms by NB4, U266 (left y-axis) and SH-SY5Y (right y-axis). Fold-changes are scaled to average. Each bar is the mean ± standard error of the mean (SEM) of technical replicates (n = 2). Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/31336622), licensed under a CC-BY license. Not internally tested by R&D Systems.

Applications for NCAM-1/CD56 Antibody

Application
Recommended Usage

CyTOF-ready

Ready to be labeled using established conjugation methods. No BSA or other carrier proteins that could interfere with conjugation.

Flow Cytometry

0.25 µg/106 cells
Sample: Human peripheral blood mononuclear cells

Immunocytochemistry

5-15 µg/mL
Sample: Immersion fixed SH-SY5Y human neuroblastoma cell line and BG01V human embryonic stem cells differentiated into neural progenitor cells

Knockout Validated

NCAM‑1/CD56 is specifically detected in U937 human histiocytic lymphoma parental cell line but is not detectable in NCAM‑1/CD56 knockout U937 human histiocytic lymphoma cell line.

Simple Western

5-25 µg/mL
Sample: Human and mouse brain (cerebellum) tissue

Western Blot

0.5 µg/mL
Sample: Human brain (cerebellum and motor cortex) tissue and mouse brain (cerebellum) tissue

Reviewed Applications

Read 2 reviews rated 5 using AF2408 in the following applications:

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Flow Cytometry

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: NCAM-1/CD56

Neural cell adhesion molecule 1 (NCAM-1) is a multifunctional member of the Ig superfamily. It belongs to a family of membrane-bound glycoproteins that are involved in Ca++ independent cell matrix and homophilic or heterophilic cell-cell interactions. NCAM-1 specifically binds to heparan sulfate proteoglycans (1), the extracellular matrix protein agrin (2), and several chondroitin sulfate proteoglycans that include neurocan and phosphocan (3). There are three main forms of human NCAM-1 that arise by alternate splicing. These are designated NCAM-120/NCAM-1 (761 amino acids [aa]), NCAM-140 (848 aa), and NCAM-180 (1120 aa). NCAM-120 is GPI-linked, while NCAM-140 and NCAM-180 are type I transmembrane glycoproteins (4‑6). Additional alternate splicing adds considerable diversity to all three forms, and extracellular proteolytic processing is possible for NCAM-180 (7‑8). NCAM-1 is synthesized as a 761 aa preproprecursor that contains a 19 aa signal sequence, a 722 aa GPI-linked mature region, and a 20 aa C-terminal prosegment (4). The molecule contains five C-2 type Ig-like domains and two fibronectin type-III domains. Human to mouse, NCAM-1 is 93% aa identical. NCAM-1 appears to be highly sialylated. The polysialyation of NCAM-1 reduces its adhesive property and increases its neurite outgrowth promoting features (9). NCAM-1 in the adult brain shows a decline of sialylation relative to earlier developmental periods. In regions that retain a high degree of neuronal plasticity, however, the adult brain continues to express polysialylation-NCAM-1, suggesting sialylation of NCAM-1 is involved in regenerative processes and synaptic plasticity (10‑13).

References

  1. Burg, M.A. et al. (1995) J. Neurosci. Res. 41:49.
  2. Storms, S.D. and U. Rutishauser (1998) J Biol. Chem. 273:27124.
  3. Margolis, R.K. et al. (1996) Perspect. Dev. Neurobiol. 3:273.
  4. Dickson, G. et al. (1987) Cell 50:1119.
  5. Lanier, L.L. et al. (1991) J. Immunol. 146:4421.
  6. Hemperly, J.J. et al. (1990) J. Mol. Neurosci. 2:71.
  7. Rutishauser, U.and C. Goridis (1986) Trends Genet. 2:72.
  8. Vawter, M.P. et al. (2001) Exp. Neurol. 172:29.
  9. Rutihauser, U. (1990) Adv. Exp. Med. Biol. 265:179.
  10. Becker, C.G. et al. (1996) J. Neurosci. Res. 45:143.
  11. Doherty, P. et al. (1995) J. Neurobiol. 26:437.
  12. Eckardt, M. et al. (2000) J. Neurosci. 20:5234.
  13. Muller, D. et al. (1996) Neuron 17:413.

Long Name

Neural Cell Adhesion Molecule

Alternate Names

CD56, NCAM1

Entrez Gene IDs

4684 (Human); 17967 (Mouse); 24586 (Rat)

Gene Symbol

NCAM1

UniProt

NP_001070150

Additional NCAM-1/CD56 Products

Product Documents for NCAM-1/CD56 Antibody

Certificate of Analysis

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Product Specific Notices for NCAM-1/CD56 Antibody

For research use only

Citations for NCAM-1/CD56 Antibody

Customer Reviews for NCAM-1/CD56 Antibody (2)

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  • Human/Mouse NCAM-1/CD56 Antibody
    Name: WonHee Kim
    Application: Western Blot
    Sample Tested: Adult brain (hippocampus), Brain olfactory membrane fraction and mouse brain
    Species: Mouse
    Verified Customer | Posted 04/15/2021
    This antibody is working great without non-specific bands.
    NCAM-1/CD56 Antibody AF2408
  • Human/Mouse NCAM-1/CD56 Antibody
    Name: Anonymous
    Application: Immunohistochemistry
    Sample Tested: Mouse brain cancer sample and Mouse brain cancer tissue
    Species: Mouse
    Verified Customer | Posted 01/19/2017
    Immunohistochemistry-Floating section

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