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. 2002 Nov 12;131(2):173–182. doi: 10.1016/0022-1759(90)90188-2

Kinetics of the in vitro antibody response to transmissible gastroenteritis (TGE) virus from pig mesenteric lymph node cells, using the ELISASPOT and ELISA tests

P Berthon 1,∗, S Bernard 1, H Salmon 1, RM Minns 2
PMCID: PMC7130707  PMID: 2167914

Abstract

A method is described for in vitro studies of viral humoral immune responses in the pig. After oral immunization with transmissible gastroenteritis (TGE) coronavirus, antibody production from primed mesenteric lymph node cells was revealed by an in vitro boost with viral antigen. For the latter the leukocytes were co-cultured with UV-inactivated virus using a variety of different methods of antigenic stimulation. Enumeration of specific antibody-secreting cells (ASC) and titration of secreted anti-virus antibodies were performed with ELISASPOT (using 3-amino 9-ethyl carbazole as the peroxidase chromogen) and ELISA tests respectively, according to the Ig isotype. The results showed a close relationship between ASC numbers and secreted antibody titres. The best in vitro antibody synthesis was observed when the sensitized cells were maintained in contact with virus during the whole culture period. Antibody responses were defined by a kinetic profile characterized by a narrow peak, with a maximum occuring after 4 and 6 days of culture and with the IgA response appearing earlier than the IgG. This methodology, which analyses specific antibody responses at the cellular level, may permit studies on the mechanisms of Ig isotype regulation. Extended to leukocytes from other organs of the immune system, it may also constitute an in vitro model to study antibody responses expressed in different lymphoid tissues of the pig.

Keywords: ELISASPOT, ELISA, Antibody-secreting cell, Mesenteric lymph node, Coronavirus, (Pig)

Abbreviations: ASC, antibody-secreting cells; ELISA, enzyme-linked immunosorbent assay; FCS, fetal calf serum; GALT, gut-associated lymphoid tissue; Ig, immunoglobulin; mAb, monoclonal antibody; MLN, mesenteric lymph node; MRBC, mouse red blood cells; NPS, normal pig serum; OD, optical density; OVA, ovalbumin; PBS, phosphate-buffered saline; p.f.u., plaque-forming units; SFC, spot-forming cells; ST, swine testis; TGE, transmissible gastroenteritis; TGEV, transmissible gastroenteritis virus

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