Optimisation of memory B cell expansion / differentiation ex vivo for interrogation of rare peripheral memory B cell subset

Human memory B cells play a vital role in the long-term : Background protection of the host from pathogenic re-challenge. In recent years the importance of a number of different memory B cell subsets that can be formed in response to vaccination or infection has started to become clear. To study memory B cell responses, cells can be cultured allowing for an ex vivo, increase in cell number and activation of these quiescent cells, providing sufficient quantities of each memory subset to enable full investigation of functionality. However, despite numerous papers being published demonstrating bulk memory B cell culture, we could find no literature on optimised conditions for the study of memory B cell subsets, such as IgM memory B cells. Following a literature review, we carried out a large screen of : Methods memory B cell expansion conditions to identify the combination that induced the highest levels of memory B cell expansion. We subsequently used a novel Design of Experiments approach to finely tune the optimal memory B cell expansion and differentiation conditions for human memory B cell subsets. Finally, we characterised the resultant memory B cell subpopulations by IgH sequencing and flow cytometry. The application of specific optimised conditions induce multiple : Results rounds of memory B cell proliferation equally across Ig isotypes, differentiation of memory B cells to antibody secreting cells, and importantly do not alter the Ig genotype of the stimulated cells. Overall, our data identify a memory B cell culture system that : Conclusions offers a robust platform for investigating the functionality of rare memory B cell subsets to infection and/or vaccination. 1 1 2 3