# Protocol for the characterization of autoproliferating B and T cells from persons with multiple sclerosis

## TL;DR

This paper provides a detailed protocol for isolating and expanding autoproliferating B and T cells from people with multiple sclerosis.

## Contribution

A new protocol is introduced for characterizing autoproliferating lymphocytes in multiple sclerosis patients.

## Key findings

- Steps for CFSE labeling and cell culture to track lymphocyte division are detailed.
- Procedures for isolating autoproliferating B and T cells are outlined.
- Methods for in vitro T cell expansion are described.

## Abstract

Autoproliferation (AP) is the spontaneous in vitro activation and proliferation of B- and T cells in the absence of exogenous stimulation. Here, we present a protocol to examine AP using peripheral blood mononuclear cells (PBMCs) derived from persons with multiple sclerosis (pwMS). We describe steps for the isolation of autoproliferating (AP+) lymphocytes as well as the expansion of AP+ T cells for downstream analyses. We detail procedures for standard fluorescence-activated cell sorting (FACS) and cell culturing.

For complete details on the use and execution of this protocol, please refer to Jelcic et al.1,2

•Steps for CFSE labeling and cell culture to track lymphocyte division•Steps for isolation of autoproliferating B and T cells for analysis•Steps for in vitro T cell expansion

Steps for CFSE labeling and cell culture to track lymphocyte division

Steps for isolation of autoproliferating B and T cells for analysis

Steps for in vitro T cell expansion

Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Autoproliferation (AP) is the spontaneous in vitro activation and proliferation of B and T cells in the absence of exogenous stimulation. Here, we present a protocol to examine AP using peripheral blood mononuclear cells (PBMCs) derived from persons with multiple sclerosis (pwMS). We describe steps for the isolation of autoproliferating (AP+) lymphocytes as well as the expansion of AP+ T cells for downstream analyses. We detail procedures for standard fluorescence-activated cell sorting (FACS) and cell culturing.