Evaluation of Cross-presentation in Bone Marrow-derived Dendritic Cells in vitro and Splenic Dendritic Cells ex vivo Using Antigen-coated Beads.
Bio Protoc
; 6(22)2016 Nov 20.
Article
en En
| MEDLINE
| ID: mdl-28239619
ABSTRACT
Antigen presentation by MHC class I molecules, also referred to as cross-presentation, elicits cytotoxic immune responses. In particular, dendritic cells (DC) are the most proficient cross-presenting cells, since they have developed unique means to control phagocytic and degradative pathways. This protocol allows the evaluation of antigen cross-presentation both in vitro (by using bone marrow-derived DC) and ex vivo (by purifying CD8+ DC from spleen after incorporation of particulate antigen) using ovalbumin (OVA)-coupled particles. Cross-presentation efficiency is measured by three different readouts the B3Z hybridoma T cell line (Karttunen et al., 1992) and stimulation of antigen-specific CD8+ T cells (OT-I) (Kurts et al., 1996), either analyzing OT-I activation by CD69 expression or OT-I proliferation after labeling them with carboxyfluorescein succinimidyl ester (CFSE). By using this approach, we could show recently that DCs are able to increase cross-presentation efficiency transiently upon engagement of TLR4 (Alloatti et al., 2015).
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MEDLINE
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En
Revista:
Bio Protoc
Año:
2016
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Article
País de afiliación:
Francia