Epitope mapping, expression and post-translational modifications of two isoforms of CD33 (CD33M and CD33m) on lymphoid and myeloid human cells

Glycobiology. 2011 Jun;21(6):757-70. doi: 10.1093/glycob/cwq220. Epub 2011 Jan 28.


We have tested the usefulness of several commercial anti-CD33 monoclonal antibodies (mAb) to determine the expression and localization of the two CD33 isoforms on several hematopoietic cell lines. The expression of the isoform CD33m, a CD33 transmembrane splice variant lacking the ligand-binding V immunoglobulin (Ig)-like domain, was detected by RT-polymerase chain reaction, western blot, confocal microscopy and flow cytometry on the membrane of several human cell types. CD33m was only detected by the anti-CD33 mAb HIM3-4 on the cell surface, whereas WM53, P67.6, 4D3, HIM3-4, WM54, D3HL60.251 or MY9 detected the CD33M isoform, indicating that HIM3-4 is the only mAb recognizing CD33 C(2) Ig domain. Accordingly, HIM3-4 binding to CD33 did not interfere with the binding of other antibodies against the CD33 V-domain. P67.6 mAb interfered with recognition by the rest of antibodies specific for the V domain. HIM3-4 staining could be increased after the sialidase treatment of all CD33(+) cells. However, this increase was stronger in activated T cells, suggesting a CD33 masking state in this cell population. Confocal microscopy analysis of CD33m HEK 293T-transfected cells revealed that this protein is expressed on the cell membrane and also detected in the Golgi compartment. CD33 is constitutively located outside the lipid raft domains, whereas cross-linked CD33 is highly recruited to this signaling platform. The unique ability of HIM3-4 mAb to detect the masking state of CD33 on different cell lineages makes it a good tool to improve the knowledge of the biological role of this sialic acid-binding Ig-like lectin.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Antibodies, Monoclonal / immunology
  • Antigen-Antibody Reactions
  • Antigens, CD / biosynthesis*
  • Antigens, CD / genetics
  • Antigens, CD / immunology*
  • Antigens, CD / metabolism
  • Antigens, Differentiation, Myelomonocytic / biosynthesis*
  • Antigens, Differentiation, Myelomonocytic / genetics
  • Antigens, Differentiation, Myelomonocytic / immunology*
  • Antigens, Differentiation, Myelomonocytic / metabolism
  • Cell Line
  • Epitope Mapping*
  • HL-60 Cells
  • Humans
  • Lymphocytes / immunology
  • Lymphocytes / metabolism*
  • Myeloid Cells / immunology
  • Myeloid Cells / metabolism*
  • Protein Isoforms / biosynthesis
  • Protein Isoforms / genetics
  • Protein Isoforms / immunology
  • Protein Isoforms / metabolism
  • Protein Processing, Post-Translational*
  • Sialic Acid Binding Ig-like Lectin 3


  • Antibodies, Monoclonal
  • Antigens, CD
  • Antigens, Differentiation, Myelomonocytic
  • CD33 protein, human
  • Protein Isoforms
  • Sialic Acid Binding Ig-like Lectin 3