Development and immunodiagnostic application of 30 kDa monoclonal antibody against coproantigens of Strongyloides in animal model
Intestinal strongyloidiasis is usually underdiagnosed and the available parasitological techniques such as stool examinations based on Baermann or agar plate cultures are time-consuming, which involved three specimens collected at the different time intervals for more accurate and sensitive appli...
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| Main Author: | |
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| Format: | Thesis |
| Language: | English |
| Published: |
2017
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| Subjects: | |
| Online Access: | http://psasir.upm.edu.my/id/eprint/92312/1/FPSK%28p%29%202018%2012%20-%20IR.pdf |
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| Summary: | Intestinal strongyloidiasis is usually underdiagnosed and the available parasitological
techniques such as stool examinations based on Baermann or agar plate cultures are
time-consuming, which involved three specimens collected at the different time
intervals for more accurate and sensitive application. The current available preferred
serological technique for diagnostic of intestinal strongyloidiasis shows the crossreaction
with other intestinal helminths. This preliminary study is aimed to produce
and evaluate monoclonal antibody against the coproantigen of Strongyloides ratti in
the animal model. Strongyloides ratti infection model was established and
maintained in immunosuppressed Sprague Dawley rats. The saline extract protein
from the infective larvae (iL3) was used as antigens for the immunisation of BALB/c
mice. The splenocytes harvested from the immunised mice were fused with
myeloma (SP2/0) cells for hybridoma production. Supernatants from the positive
hybrids were screened by indirect ELISA. The purified IgG2b MAb was
characterised by western blots and evaluated in sandwich-ELISA for reactivity
against the homologous and heterologous antigens which include Toxocara canis,
Toxocara cati, Ancylostoma caninum and Ascaris suum. An IgG2b MAb that
recognises 30 kDa molecular weight proteins associated with strongyloidiasis and a
cross-reaction with a 30 kDa for Ancylostoma caninum were observed. The IgG2b
MAb was recognised by two Strongyloides ratti antigens and saline extract antigen
of Ancylostoma caninum but did not react with other heterologous antigens in both
assays. The antigen detection limit by sandwich ELISA was 75 ng/mL. Seventeen
out of the twenty strongyloidiasis experimentally infected rat faeces (85%) evaluated
for coproantigen using the IgG2b MAb have shown antigen-positive reactions in
sandwich-ELISA. Similarly, only Ancylostoma caninum from dog faeces were
reactive against the IgG2b MAb from all samples tested for cross-reactivity. This study concluded that the IgG2b MAb produced was able to detect Strongyloidiasis
and Ancylostomiasis in animal models. |
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