TY - JOUR
T1 - Use of fluorescent immuno-chemistry for the detection of Edwardsiella ictaluri in channel catfish (I. punctatus) samples
AU - Menanteau-Ledouble, Simon
AU - Lawrence, Mark
PY - 2011/5
Y1 - 2011/5
N2 - While Edwardsiella ictaluri is a major pathogen of channel catfish Ictalurus punctatus and has been discovered nearly three decades ago 1,2, so far, to the best of these authors' knowledge, no method has been developed to allow for the in situ visualization of the bacteria in histological sections. While bacterial localization has been determined in vivo in previous studies using plate counts 3, radiometric labeled 4, or bioluminescent bacteria 5, most of these studies have only been performed at the gross organ level, with one exception 6. This limitation is of particular concern because E. ictaluri has a complex infection cycle 1,7, and it has a variety of virulence factors 8,9. The complex interaction of E. ictaluri with its host is similar in many respects to Salmonella typhi 10, which is in the same taxonomic family. Here we describe a technique allowing for the detection of bacteria using indirect immuno-histochemistry using the monoclonal Ed9 antibody described by Ainsworth et al.11. Briefly, a blocking serum is applied to paraffin embedded histological sections to prevent non-specific biding. Then, the sections are incubated with the primary antibody: E. ictaluri specific monoclonal antibody Ed9. Excess antibodies are rinsed away and the FitC labeled secondary antibodies are added. After rinsing, the sections are mounted with a fluorescent specific mounting medium. This allowed for the detection of E. ictaluri in situ in histological sections of channel catfish tissues.
AB - While Edwardsiella ictaluri is a major pathogen of channel catfish Ictalurus punctatus and has been discovered nearly three decades ago 1,2, so far, to the best of these authors' knowledge, no method has been developed to allow for the in situ visualization of the bacteria in histological sections. While bacterial localization has been determined in vivo in previous studies using plate counts 3, radiometric labeled 4, or bioluminescent bacteria 5, most of these studies have only been performed at the gross organ level, with one exception 6. This limitation is of particular concern because E. ictaluri has a complex infection cycle 1,7, and it has a variety of virulence factors 8,9. The complex interaction of E. ictaluri with its host is similar in many respects to Salmonella typhi 10, which is in the same taxonomic family. Here we describe a technique allowing for the detection of bacteria using indirect immuno-histochemistry using the monoclonal Ed9 antibody described by Ainsworth et al.11. Briefly, a blocking serum is applied to paraffin embedded histological sections to prevent non-specific biding. Then, the sections are incubated with the primary antibody: E. ictaluri specific monoclonal antibody Ed9. Excess antibodies are rinsed away and the FitC labeled secondary antibodies are added. After rinsing, the sections are mounted with a fluorescent specific mounting medium. This allowed for the detection of E. ictaluri in situ in histological sections of channel catfish tissues.
KW - Catfish
KW - Edwardsiella ictaluri
KW - Enteric septicemia of channel catfish
KW - Fish
KW - Histology
KW - Ictalurus punctatus
KW - Immunohistochemistry
KW - Immunology
KW - Issue 51
UR - http://www.scopus.com/inward/record.url?scp=80355139306&partnerID=8YFLogxK
U2 - 10.3791/2687
DO - 10.3791/2687
M3 - Journal article
C2 - 21610669
AN - SCOPUS:80355139306
SN - 1940-087X
JO - Journal of Visualized Experiments
JF - Journal of Visualized Experiments
IS - 51
M1 - e2687
ER -