B Cell Activation on an Antigen-Coated Coverslip

Published: August 31, 2023

Abstract

Source: Ibañez-Vega, J., et al., Studying Organelle Dynamics in B Cells During Immune Synapse Formation. J. Vis. Exp. (2019)

This video demonstrates B cell activation in vitro. The process involves antigen interaction with B cell receptors, leading to cytoskeleton remodeling, immune synapse formation, lysosome movement, antigen internalization, processing, and subsequent presentation on MHC molecules for B cell activation.

Protocol

1. Preparation of poly-L-lysine coverslips Before the B cell activation assay with Ag-coated beads, prepare poly-L-lysine-coated coverslips (PLL-coverslips). Use a 50 mL tube containing 40 mL of 0.01% w/v of PLL solution and immerse the 12 mm-diameter coverslips into the solution. Rotate overnight at RT. Wash the coverslips with 1xPBS and leave to dry on a 24-well plate lid covered with paraffin film. Proceed with B cell activation. 2. B cell…

Disclosures

The authors have nothing to disclose.

Materials

IIA1.6 (A20 variant) mouse B-lymphoma cells ATCC TIB-208 Murine B-cell lymphoma of Balb/c origin that expresses an IgG-containing BCR on its surface without FcγIIR
CaCl2 Winkler CA-0520
Glycine Winkler  BM-0820
HyClone Fetal bovine serum Thermo Fisher Scientific SH30071.03 Heat inactivate at 56 C for 30 min
KCl Winkler PO-1260
NaCl Winkler SO-1455
Parafilm M P1150-2
Penicillin-Streptomycin Thermo Fisher Scientific 15140122 Liquid
Poly-L-Lysine Sigma P8920 Dilute with sterile water
RPMI-1640 Biological Industries 01-104-1A
Sodium pyruvate Thermo Fisher Scientific 11360070
Tube 50 ml Corning 353043
2-mercaptoethanol Thermo Fisher Scientific 21985023
Glutamine Thermo Fisher Scientific 35050061
Goat-anti-mouse IgG antibody Jackson ImmunoResearch 315-005-003 IIA1.6 positive ligand

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Cite This Article
B Cell Activation on an Antigen-Coated Coverslip. J. Vis. Exp. (Pending Publication), e21611, doi: (2023).

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