Mapping of the continuous epitopes displayed on the Clostridium perfringens type D epsilon-toxin

Alves, Guilherme Guerra; Machado-de-Ávila, Ricardo Andrez; Chávez-Olórtegui, Carlos; Silva, Rodrigo; Lobato, Francisco Carlos Faria

doi:10.1016/j.bjm.2016.10.023

Cited by 4 publications

(3 citation statements)

References 26 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Two of these neutralizing antibodies (PεTX6 and PεTX9) recognize the same linear epitope (89-LLTNDTQQ-96) located in the β-sandwich domain III of the protein [ 33 ] at the C-terminal extremity of the four-stranded sheet and its following loop just before the final three-stranded sheet. To our knowledge, this epitope was never previously identified as important for cytotoxicity [ 36 ]. Concentrations of our neutralizing antibodies necessary to maintain 50% cell viability were determined to be in the 1 to 10 μg/mL range, i.e.…”

Section: Discussionmentioning

confidence: 99%

Highly sensitive sandwich immunoassay and immunochromatographic test for the detection of Clostridial epsilon toxin in complex matrices

et al. 2017

View full text Add to dashboard Cite

Epsilon toxin is one of the four major toxins of Clostridium perfringens. It is the third most potent clostridial toxin after botulinum and tetanus toxins and is thus considered as a potential biological weapon classified as category B by the Centers for Disease Control and Prevention (CDC). In the case of a bioterrorist attack, there will be a need for a rapid, sensitive and specific detection method to monitor food and water contamination by this toxin, and for a simple human diagnostic test. We have produced and characterized five monoclonal antibodies against common epitopes of epsilon toxin and prototoxin. Three of them neutralize the cytotoxic effects of epsilon toxin in vitro. With these antibodies, we have developed highly sensitive tests, overnight and 4-h sandwich enzyme immunoassays and an immunochromatographic test performed in 20 min, reaching detection limits of at least 5 pg/mL (0.15 pM), 30 pg/mL (0.9 pM) and 100 pg/mL (3.5 pM) in buffer, respectively. These tests were also evaluated for detection of epsilon toxin in different matrices: milk and tap water for biological threat detection, serum, stool and intestinal content for human or veterinary diagnostic purposes. Detection limits in these complex matrices were at least 5-fold better than those described in the literature (around 1 to 5 ng/mL), reaching 10 to 300 pg/mL using the enzyme immunoassay and 100 to 2000 pg/mL using the immunochromatographic test.

show abstract

Section: Discussionmentioning

confidence: 99%

Highly sensitive sandwich immunoassay and immunochromatographic test for the detection of Clostridial epsilon toxin in complex matrices

et al. 2017

View full text Add to dashboard Cite

show abstract

“…It is difficult to predict which epitopes the non-neutralizing antibodies recognize. By using overlapping ETX peptides, Alves et al were able to determine 15 epitopes that are recognized in ETX hyperimmune rabbit serum and identified four epitopes that appear to be immunodominant [34]. Interestingly, the most immunodominant peptide was ( 116 TTTHTVGTSIQATAKFTVPFN 136 ).…”

Section: Discussionmentioning

confidence: 99%

“…Domain III, which contains the C-terminus, is also important in membrane insertion and oligomerization as mutations in domain III block ETX oligomerization [23,30]. As suggested by previous experiments [34,35], it is plausible that antibodies directed against external epitopes in any of ETX’s three domains could neutralize cytotoxicity either by blocking ETX binding or oligomerization and pore formation.…”

Section: Introductionmentioning

confidence: 99%

A Novel Panel of Rabbit Monoclonal Antibodies and Their Diverse Applications Including Inhibition of Clostridium perfringens Epsilon Toxin Oligomerization

et al. 2018

View full text Add to dashboard Cite

The pore-forming epsilon toxin (ETX) produced by Clostridium perfringens is among the most lethal bacterial toxins known. Sensitive antibody-based reagents are needed to detect toxin, distinguish mechanisms of cell death, and prevent ETX toxicity. Using B-cell immuno-panning and cloning techniques, seven ETX-specific monoclonal antibodies were generated from immunized rabbits. ETX specificity and sensitivity were evaluated via western blot, ELISA, immunocytochemistry (ICC), and flow cytometry. ETX-neutralizing function was evaluated both in vitro and in vivo. All antibodies recognized both purified ETX and epsilon protoxin via western blot with two capable of detecting the ETX-oligomer complex. Four antibodies detected ETX via ELISA and three detected ETX bound to cells via ICC or flow cytometry. Several antibodies prevented ETX-induced cell death by either preventing ETX binding or by blocking ETX oligomerization. Antibodies that blocked ETX oligomerization inhibited ETX endocytosis and cellular vacuolation. Importantly, one of the oligomerization-blocking antibodies was able to protect against ETX-induced death post-ETX exposure in vitro and in vivo. Here we describe the production of a panel of rabbit monoclonal anti-ETX antibodies and their use in various biological assays. Antibodies possessing differential specificity to ETX in particular conformations will aid in the mechanistic studies of ETX cytotoxicity, while those with ETX-neutralizing function may be useful in preventing ETX-mediated mortality.

show abstract

Harnessing an Integrative In Silico Approach to Engage Highly Immunogenic Peptides in an Antigen Design Against Epsilon Toxin (ETX) of Clostridium perfringens

Mahboobi

Sedighian

Malekara

et al. 2020

Int J Pept Res Ther

View full text Add to dashboard Cite

Mapping of the continuous epitopes displayed on the Clostridium perfringens type D epsilon-toxin

Cited by 4 publications

References 26 publications

Highly sensitive sandwich immunoassay and immunochromatographic test for the detection of Clostridial epsilon toxin in complex matrices

Highly sensitive sandwich immunoassay and immunochromatographic test for the detection of Clostridial epsilon toxin in complex matrices

A Novel Panel of Rabbit Monoclonal Antibodies and Their Diverse Applications Including Inhibition of Clostridium perfringens Epsilon Toxin Oligomerization

Harnessing an Integrative In Silico Approach to Engage Highly Immunogenic Peptides in an Antigen Design Against Epsilon Toxin (ETX) of Clostridium perfringens

Contact Info

Product

Resources

About