Human CD99 Antibody Summary
Accession # P14209
Please Note: Optimal dilutions should be determined by each laboratory for each application. General Protocols are available in the Technical Information section on our website.
Detection of Human CD99 by Western Blot. Western blot shows lysates of NCI-H460 human large cell lung carcinoma cell line, U251-MG human malignant glioblastoma cell line, amd SK-Mel-28 human malignant melanoma cell line. PVDF membrane was probed with 0.25 µg/mL of Goat Anti-Human CD99 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF3968) followed by HRP-conjugated Anti-Goat IgG Secondary Antibody (Catalog # HAF017). A specific band was detected for CD99 at approximately 32 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.
Detection of CD99 in Human CD3+Lymphocytes by Flow Cytometry. Human whole blood CD3+lymphocytes were stained with Goat Anti-Human CD99 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF3968, filled histogram) or control antibody (Catalog # AB-108-C, open histogram), followed by Phycoerythrin-conjugated Anti-Goat IgG Secondary Antibody (Catalog # F0107).
CD99 in HepG2 Human Cell Line. CD99 was detected in immersion fixed HepG2 human hepatocellular carcinoma cell line using Goat Anti-Human CD99 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF3968) 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 cell membranes. View our protocol for Fluorescent ICC Staining of Cells on Coverslips.
Detection of Human CD99 by Simple WesternTM. Simple Western lane view shows lysates of MOLT-4 human acute lymphoblastic leukemia cell line, loaded at 0.2 mg/mL. A specific band was detected for CD99 at approximately 37 kDa (as indicated) using 20 µg/mL of Goat Anti-Human CD99 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF3968) followed by 1:50 dilution of HRP-conjugated Anti-Goat IgG Secondary Antibody (Catalog # HAF019). This experiment was conducted under reducing conditions and using the 12-230 kDa separation system.
Preparation and Storage
- 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.
CD99 (also named MIC2, E2 and thymic leukemia antigen) is the founding member of the CD99 family of molecules. The CD99 family contains four members; CD99, CD99L2, XG and the pseudogene CD99L1 (1, 2, 3). Native human CD99 is 32 kDa in size and exists as a type I transmembrane glycoprotein. This is referred to as the long, or type I isoform. It is synthesized as a 185 amino acid (aa) precursor that contains a 22 aa signal sequence, a 100 aa extracellular domain (ECD), a 25 aa transmembrane segment, and a 38 aa cytoplasmic region (4). The ECD contains no identifiable motifs, N‑linked glycosylation sites, or cysteine residues; it does possess sites for O-linked glycosylation. The cytoplasmic region, albeit short, does have signal transduction capability (5). There are apparently multiple isoforms for human CD99. One shows a 16 aa deletion in the ECD (aa 34‑49), a second shows a 38 aa deletion in the cytoplasmic region (aa 122‑159), and a third exhibits a three aa truncation at the C-terminus (6, 7, 8). The best studied isoform shows an Asp‑Gly substitution for the C‑terminal 27 amino acids. This is referred to as the 28 kDa type II isoform (9). The type I and II isoforms have distinctive signal transduction pathways (FAK-src for type I; PI3K plus src-ERK1/2 for type II), and mediate clearly different biological outcomes (5, 9, 10). The two numbered isoforms may or may not co‑exist on the same cells. Peripheral T cells have only the long isoform, while double-positive thymocytes express both isotypes. What is unclear is the monomeric vs. dimeric status of CD99. In mouse, CD99 reportedly forms disulfide-linked homodimers (11). In human, however, CD99 is reportedly monomeric if only a type I isoform, and a covalent heterodimer if co‑expressing type I and II isoforms (12, 13). Cells known to express CD99 include fibroblasts, neutrophils, T cells, double-positive thymocytes, CD34+ stem cells, monocytes and endothelial cells (2, 12, 14, 15). Homophilic interaction between CD99 on the neutrophil and CD99 on the endothelial cell regulates the transendothelial migration of neutrophils during inflammation (16). Human CD99 is only 48% aa identical to mouse CD99 (17).
- Wilson, M.D. et al. (2006) Physiol Genomics 27:201.
- Petri, B. and M.G. Bixel (2006) FEBS J. 273:4399.
- Suh, Y.H. et al. (2003) Gene 307:63.
- Gelin, C. et al. (1989) EMBO J. 8:3253.
- Byun, H-J. et al. (2006) J. Biol. Chem. 281:34833.
- GenBank Accession # EAW98698.
- GenBank Accession # EAW98699.
- GenBank Accession # EAW98700.
- Hahn, H-J. et al. (1997) J. Immunol. 159:2250.
- Scotlandi, K. et al. (2007) Oncogene Apr 30; [Epub ahead of print].
- Park, S.H. et al. (2005) Gene 353:177.
- Schenkel, A.R. et al. (2002) Nat. Immunol. 3:143.
- Alberti, I. et al. (2002) FASEB J. 16:1946.
- Imbert, A-M. et al. (2006) Blood 108:2578.
- Dworzak, M.N. et al. (1994) Blood 83:415.
- Lou, O. et al. (2007) J. Immunol. 178:1136
- Shiratori, I. et al. (2004) J. Exp. Med. 199:525.
Citations for Human CD99 Antibody
R&D Systems personnel manually curate a database that contains references using R&D Systems products. The data collected includes not only links to publications in PubMed, but also provides information about sample types, species, and experimental conditions.
Citations: Showing 1 - 2
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Ectodomain shedding of CD99 within highly conserved regions is mediated by the metalloprotease meprin ? and promotes transendothelial cell migration
Authors: Tillmann Bedau
FASEB J, 2016;0(0):.
Sample Types: Cell Lysates
Applications: Western Blot
Evolutionarily conserved paired immunoglobulin-like receptor alpha (PILRalpha) domain mediates its interaction with diverse sialylated ligands.
Authors: Sun Y, Senger K, Baginski TK, Mazloom A, Chinn Y, Pantua H, Hamidzadeh K, Ramani SR, Luis E, Tom I, Sebrell A, Quinones G, Ma Y, Mukhyala K, Sai T, Ding J, Haley B, Shadnia H, Kapadia SB, Gonzalez LC, Hass PE, Zarrin AA
J. Biol. Chem., 2012;287(19):15837-50.
Sample Types: Whole Cells
Applications: Flow Cytometry
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