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A monoclonal antibody monitoring band 3 modifications in human red blood cells

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Abstract

Morphologic and methabolic erythrocyte modifications are thought to be the basis of cell removal from circulating blood. A significant role has been ascribed to the immunological network which may remove aged or misshapen erythrocytes through the binding of specific autoantibodies. Along this line recent observations indicate that a senescence antigen appears in consequence of postsynthetic modifications of band 3, one of the most important erythrocyte membrane proteins, which accounts for many functional activities of the red cells. On this basis, we raised a mouse hybridoma anti-band 3 monoclonal antibody (B6 MoAb) of the IgG2a class which monitors band 3 differences among normal red blood cells separated by Percoll density gradient. These differences are outlined by the decrease of B6 MoAb binding to band 3 monomer, the appearance of an 80–90 kDa new band, lighter than band 3, and the increase of low molecular weight fragments in the 4.5 region. The B6 MoAb appears to be very useful in detecting modifications of band 3 since it bind to a 19 kDa Chy-Try fragment estimated to be sensitive to aging.

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Abbreviations

PBS:

Phosphate Buffer Saline

MoAb:

Monoclonal Antibody

RBCs:

Red Blood Cells

PMSF:

Phenylmethylsulphonyl Fluoride

PVC:

Polyvinyl Chloride

ACD:

Acid Citrate Dextrose

HMWP:

High Molecular Weight Polymers

Chy-Try:

Chymotrypsin-Trypsin Digested

i.p.:

intraperitoneum

ELISA:

Enzyme Linked Immuno Sorbent Assay

Hepes:

4-(2-Hydroxyethyl)-piperazine-1-ethane-sulfonic acid. Enzymes: trypsin (EC 3.4.21.4), chymotrypsin (EC 3.4.21.1), neuraminidase (EC 3.2.1.18)

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Authors and Affiliations

  1. Laboratorio di Immunologia, Laboratorio di Biochimica Clinica, Istituto Superiore di Sanità, Roma, Italy

    Lucietta Ferroni & Patrizia Caprari

  2. Dipartimento di Medicina Sperimentale e Scienze Biochimiche, Università ‘Tor Vergata’, Roma, Italy

    Anna Giuliani, Stefano Marini, Saverio G. Condò & Bruno Giardina

  3. Istituto di Ricerche sulla Senescenza, Sigma Tau, Pomezia, Italy

    Maria T. Ramacci

Authors
  1. Anna Giuliani
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  2. Stefano Marini
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  3. Lucietta Ferroni
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  4. Patrizia Caprari
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  5. Saverio G. Condò
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  6. Maria T. Ramacci
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  7. Bruno Giardina
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Giuliani, A., Marini, S., Ferroni, L. et al. A monoclonal antibody monitoring band 3 modifications in human red blood cells. Mol Cell Biochem 117, 43–51 (1992). https://doi.org/10.1007/BF00230409

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  • DOI: https://doi.org/10.1007/BF00230409

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