title: Characterization of local antibody responses to the gastrointestinal parasite Haemonchus contortus.
abstract: The ability to identify antigens associated with an infection has generally relied on the use of serum antibodies produced by infected or previously exposed individuals. A major drawback with the use of serum is that it does not necessarily reflect the local antibody response at mucosal tissue sites. This study describes an approach that allows the use of antibodies generated close to the infection site to detect the transient expression of stage-specific antigens during infection with the gastrointestinal parasite Haemonchus contortus. This was achieved by infecting immune sheep with H. contortus larvae and removing the abomasal lymph nodes draining the infection site shortly after the challenge infection. Antibody-secreting cell (ASC) probes were generated from these lymph nodes after short-term in vitro culture of cell suspensions, which allowed the accumulation of antibodies secreted by in vivo-induced ASC into the culture supernatant. Lymph node culture supernatants (= ASC probes) from immune sheep challenged 5 days previously were used to probe Western blots of third and fourth stage larval preparations, and revealed distinct reactivity to larval antigens. No antibody reactivity to larval antigen preparations was detected in sheep that were not challenged. The number of antigens identified using ASC probes was significantly restricted compared to either pre- or post-challenge sera. In contrast to the variability of the serum response, the specificity of ASC probes was highly repeatable between different sheep. ASC probes were also used to purify a H. contortus larval antigen by affinity chromatography, which allowed limited biochemical studies to be undertaken. The antigen(s) recognized by the ASC probes were shown to be expressed on the surface of the larvae. These studies illustrate the use of a novel means of studying the local antibody response close to a mucosal infection site in order to identify and isolate stage-specific antigens expressed during infection.
authors.affiliation: Centre for Animal Biotechnology, School of Veterinary Science, University of Melbourne, Parkville, Victoria, Australia.
authors.forename: V M
authors.initials: VM
authors.lastname: Bowles
authors.forename: M R
authors.initials: MR
authors.lastname: Brandon
authors.forename: E
authors.initials: E
authors.lastname: Meeusen
mesh_terms: D000818:Animals; D000909:Antibodies, Helminth / Q000096:biosynthesis*; D000921:Antibody-Producing Cells / Q000276:immunology*; D000947:Antigens, Helminth / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification; D015153:Blotting, Western; D002846:Chromatography, Affinity; D006188:Haemonchiasis / Q000276:immunology / Q000662:veterinary*; D006189:Haemonchus / Q000276:immunology*; D007814:Larva / Q000276:immunology; D012756:Sheep; D012757:Sheep Diseases / Q000276:immunology*
chemical_list: D000909:Antibodies, Helminth; D000947:Antigens, Helminth