Characterization of rat HSA/CD24 protein bearing the B lineage-restricted epitope recognized by MAb HIS50

Hybridoma. 1996 Feb;15(1):33-42. doi: 10.1089/hyb.1996.15.33.

Abstract

HIS50 MAb recognizes a GPI-linked molecule on the surface of rat cells committed to the B lineage but is absent from cells of other hematopoietic lineages. Eukaryotic expression cloning of the cDNA has revealed that HIS50 recognizes a rat homologue of human CD24 and murine HSA. CD24/HSA exists in multiple glycoforms and plays an important role in hematopoietic and neural cell development. HIS50 is significant in that it is the only, presently available, anti-rat CD24/HSA MAb. Furthermore it is the only reported MAb with specificity restricted to B cell forms of CD24. Here we report Western blot analysis of HIS50 antigen (ag). In rat bone marrow HIS50 MAb recognized MW species in the range of 35-70 kDa. These species were shown by immunomagnetic cell sorting to be all derived from the HIS50+ cell subset. In other tissues with varying HIS50 levels as judged by immunostaining, apparent levels of HIS50 ag in the 35-70 kDa range varied accordingly. Thus, Western blot data corresponded to the immunostaining data of HIS50 ag, and the size distribution of B-restricted forms of rat CD24 appeared as wide as that reported for the more extensively expressed forms of human and murine CD24. N-Deglycosylation of cell lysates from lymphoid organs reduced the signal in the 35-70 kDa range, without appearance of lower MW HIS50-reactive species. Thus, unlike certain epitopes on human and murine CD24, HIS50 epitopes appeared (partially) N-linked carbohydrate dependent. The data reported here provide a basis for the further use of HIS50 MAb in studying the role of the highly heterogeneous CD24 molecules in cell development.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amidohydrolases
  • Animals
  • Antibodies, Monoclonal / chemistry*
  • Antibody Specificity
  • Antigen-Antibody Reactions
  • Antigens, CD / chemistry*
  • Antigens, CD / drug effects
  • Antigens, CD / immunology
  • B-Lymphocytes / immunology*
  • Blotting, Western
  • Bone Marrow Cells
  • CD24 Antigen
  • Cell Differentiation / immunology
  • Cell Separation
  • Detergents
  • Epitopes / chemistry*
  • Epitopes / immunology
  • Glycosylation
  • Male
  • Membrane Glycoproteins*
  • Mice
  • Mice, Inbred BALB C
  • Molecular Weight
  • Octoxynol
  • Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase
  • Polyethylene Glycols
  • Rats
  • Rats, Inbred F344
  • Sodium Hydroxide

Substances

  • Antibodies, Monoclonal
  • Antigens, CD
  • CD24 Antigen
  • Cd24a protein, mouse
  • Detergents
  • Epitopes
  • Membrane Glycoproteins
  • Polyethylene Glycols
  • Sodium Hydroxide
  • Octoxynol
  • Nonidet P-40
  • Amidohydrolases
  • Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase