Study of the reactivity of a monoclonal antibody that recognises human marginal zone B cells and a subset of germinal centre cells in tissue sections

• Main Author: Alyafei, Saud Abdalla Mubarak Mohamed
• Other Authors: Spencer, Jo Michele
• Published: King's College London (University of London) 2018
• Subjects: 616
• Online Access: https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.762359

Marginal Zone (MZ) cells are a subset of B cells. They reside the marginal zone outside the mantle zone of the spleen and circulate the blood in humans. They have also been identified in the crypt epithelium of tonsil, the sub-capsular sinus in lymph node and in the dome area of Peyer’s patches in gut-associated lymphoid tissue of human intestine. A monoclonal antibody (4D12) that recognises an antigen on MZ B cells and a subset of GC cells in tissue sections has been described previously but this has not been investigated in detail and features of the 4D12 antigen and the lymphocytes that express it are unknown. The re-establishment and culture of the 4D12 MoAb secreting clone after a long-term storage is described. A method to analyse the distribution of 4D12 antigen on subsets of B cells was devised. Subsets expressing 4D12 antigen included CD27+IgM+IgD+ cells, CD27+IgM+IgD- cells, transitional B cells and plasmablasts. Lower expression by mature naïve and class switched memory B cells was observed. 4D12 MoAb tended to recognise a greater proportion of CD27+IgM+IgD+ cells and CD27+IgM+IgD- cells in spleen compared to tonsil and blood. Intracellular staining showed that the 4D12 antigen is proportionally more presented in the cytoplasm than the cell surface and that cytoplasmic 4D12 antigen expression is not restricted to B cells. The identity of the antigen was sought by western blotting and mass spectrometry (by collaborators) and the output of the mass spectrometry was provided for this study. Mass spectrometry data was analysed here and a list of candidates for the identity of 4D12 antigen were selected and tested by flow cytometry.