• Parasites, Hosts and Diseases
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• v.41 no.1
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• pp.35-39
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• 2003

Although stool examination is the standard diagnostic method of clonorchiasis, serodiagnosis by ELISA using crude antigen is now widely used because of its convenience. However, ELISA diagnosis still suffers from cross-reactions, and therefore there is a need to improve the present conventional ELISA. The present study was undertaken to evaluate the diagnostic value of ELISA using excretory-secretory antigen (ESA) instead of crude antigen (CA) of Clonorchis sinensis. The diagnostic sensitivity of ELISA using excretory-secretory antigen was 92.5%, which was higher than that of ELISA using crude Clonorchis sinensis antigen (88.2%). In addition, the specificity of excretory-secretory antigen was found 93.1% while that of crude antigen was 87.8%. In summary, Clonorchis sinensis ESA was found to be a better serodiagnostic antigen than CA for ELISA.

• Parasites, Hosts and Diseases
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• v.24 no.2
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• pp.187-193
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• 1986

Enzyme-linked immunosorbent assay(ELISA) using crude and affinity-purified antigens of adult worms of Paragonimus westermani was performed for infected cat sera with different worm burden, from preinfection to 18th week after infection. Crude antigen was used with supernatant of homogenated worms by freezing-thawing method, and the supernate was centrifuged for 1 hour at 10,000 rpm at $4^{\circ}C$. Affinity-purified antigen(antibody-bound antigen) was prepared from fractions(bound and unbound) of crude antigen by affinity chromatography on CNBr-activated sepharose 4B, and IgG as a ligand was prepared from paragonimiasis cat serum(6 months infected) obtained by ammonium sulfate ($40%{\sim}45%$ saturated) precipitation method. By SDS-PAGE, crude antigen showed 22 polypeptide fractions while purified antigen showed 4 fractions: 36, 400, 34, 700, 27, 600 and 11, 500 in molecular weights. All cats were divided into five groups($G_1-G_5$) by different worm burdens. The mean of recovered worms(${\pm}SD$) and the number of cats in each group are as follows: $G_1$, 2 worms(0) and 4 cats; $G_2$, 4.75 (${\pm}0.66$) and eight; $G_3$, 10.75(${\pm}1.92$) and four; $G_4$, 23.20(${\pm}3.43$) and five; $G_5$, 48(${\pm}12.63$) and five cats. The results were summarized as follows: 1. The antibody levels(OD value) increased by worm burden in $G_1$ to $G_4$ generally. However, individual antibody levels were not exactly related with worm burden in all groups, especially there was a wide difference in $G_4$ and $G_5$. These results suggested that the worm burden in $G_4$ (about $20{\sim}30$ worms) is enough to produce antibody maximum in cats of $2{\sim}3kg$ weight. 2. The antibody levels increased significantly(p<0.05) compared to control sera at the 3rd week in $G_1$ and $G_2$, at the 2nd week in $G_3$, and at the 1st week in $G_4$ and $G_5$. Especially in the 4th week, OD value increased more in $G_1$(p<0.01) and in $G_2$ to $G_5$(p<0.001). In the pattern of antibody levels by ELISA in each group, OD in $G_1$ increased to the 18th week continuously, in $G_2$ OD was maintained same after the 16th week, but in $G_3$ it decreased after the 16th week, and it was maintained same in $G_4$ and $G_5$ after the 14th week. 3. The antibody levels by ELISA with the affinity-purified antigen were higher than those with crude antigen in all groups generally. Especially, the difference of OD values between two antigens was larger from the 4th to the 10th week. In $G_1$ and $G_2$ OD with purified antigen was higher than that with crude one to the 18th week. It was also higher in $G_3$ than that with crude antigen to the 16th week and OD of $G_4$ and $G_5$ were higher before the 14th week than that with crude antigen, however became lower at the 16th week. Consequently, the antibody level in ELISA with affinity-purified antigen was more sensitive at the early weeks after infection and in light infection groups than that with crude antigen.

• Parasites, Hosts and Diseases
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• v.24 no.2
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• pp.171-176
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• 1986

In order to obtain the most specific and sensitive antigen from crude antigens of Fasciola hepatica for the immunodiagnosis of bovine fascioliasis by the enzyme linked immunosorbent assay (ELISA), phosphate buffered saline extract of F. hepatica was prepared. The crude extract was fractionated into 7 antigens using sephadex G-100 column chromatography. Seven fractionated antigens were applied to ELISA, precipitation test and intradermal test, respectively. Results obtained are as follows: 1. The specificity (95% confidence interval in negative sera of bovine fascioliasis; Mean+$2{\times}SD$ of absorbence) of the first (MW>150,000) and the second antigens(MW 120,000) were 93.7%, but those of others including crude antigen showed 100%. 2. The sensitivity(positive sera of bovine fascioliasis having higher values with compared to the criterion) of the first, the sixth (MW 16,000) and the seventh antigen (MW<5,000) were 91.6%, 87.5% and 0%, respectively, but those of others showed all 100%. 3. The absorbance by ELISA using the fifth antigen (MW 26,000) was 8. 43-folds higher in the positive sera than that in the negative sera. This could be used as one of the most specific antigens for the immunodiagnosis of bovine fascioliasis. 4. In Ouchterlony test, precipitin lines were not found in the sera naturally infected with F. hepatica, but some were found in the sera of rabbits immunized with the crude antigens. The numbers of precipitin lines in the sera of rabbits were different in the different fractionated antigens. They were 6 in the crude, 2 in the second and the third antigens, 1 in the fourth, the fifth and the sixth antigens and absent in the seventh antigen, respectively. 5. The wheal size for bovine infected with F. hepatica was $2.46{\pm}0.15cm$ in the intradermal test antigen(saline extract of F. hepatica) supplied by the Veterinary Research Institute, Rural Development Administration, Korea. The wheal size of the first, the second and the third antigens were larger than that. of intradermal test antigen, whereas those of the fouth, the fifth, the sixth and the seventh antigens showed smaller than that of the intradermal test antigen. The results suggest that the fifth antigen may be specific antigen for the immunodiagnosis of bovine fascioliasis.

• Journal of Life Science
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• v.18 no.2
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• pp.241-248
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• 2008

This study has been carried out to secretion antibodies for the purpose of preventing the infection of Helicobacter pylori and using them as a supplement for treatment. This experiments have been separated antigens from H. pylori and observed into antibody production and the agglutination of H. pylori for the separated antigens. As major antigenic proteins separated from H. pylori, the following could be verified: 12 kinds of band for whole cell (WC), seven kinds of band for outer membrane protein (OMP), three kinds of band for crude urease, and one kind of band for lipopolysaccharide (LPS). The IgG anti-H. pylori antibody of separated antigens showed $77.9{\pm}6.4{\mu}g/ml$ for we (L), $84.9{\pm}6.4{\mu}g/ml$ for OMP, and $123.8{\pm}2.9{\mu}g/ml$ for crude urease, at the same antigen concentration of $20{\mu}g/100ull$, which showed the most at the crude urease. And it turned out that the IgA antibodies were generated with $2.5{\pm}0.32{\mu}g/ml$ for WC (L), $2.0{\pm}0.43{\mu}g/ml$ for OMP, and $1.3{\pm}0.25{\mu}g/ml$ for crude urease, which demonstrated the most for WC (L) antigens. As a result of verifying the immunogenecity of antigenic protein through the Western blotting, major antigenic substances could be confirmed as follows: 10 kinds for WC, six kinds for OMP and three kinds for crude urease. The agglutination values on the H. pylori of the antibody were $2^5,\;2^5,\;2^6\;and\;2^7$ at the antigen serums of anti-WC (H), anti-WC (L), anti-OMP and anti-crude urease, respectively, which indicated the highest for the antigen serum of anti-crude urease. The urease activation-inhibiting absorbance of antigen serum created by each antigen was $0.14{\pm}0.01$ for WC (H), $0.16{\pm}0.01$ for WC (L), $0.18{\pm}0.03$ for OMP, and $0.18{\pm}0.04$ for urease, demonstrating a significant inhibiting effect, compared with $0.26{\pm}0.02$ of the control group.

• Biomedical Science Letters
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• v.3 no.2
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• pp.89-94
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• 1997

The authors characterized the proteins of the crude antigen obtained from Clonorchis sinensis worm and excretory-secretory and billis from rabbits, experimentally infected for 3 months. Protein composition was observed after adding a cysteine proteinase inhibitor E-64 and a serine proteinase inhibitor PMSF, respectively. SDS-PAGE of the crude antigen from C. sinensis recovered from the infected rabbits, the crude antigen from the adult worm excretory-secretory, and the crude antigen from billis of the rabbits resolved 26, 27 and 19 profiles between 200-9 kDa, respectively. When E-64 supplemented 29, and 22 bands, respectively. More study should be carried out in the future on the immunological characteristics and the monoclonal antibody of the each antigen.

• Parasites, Hosts and Diseases
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• v.39 no.2
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• pp.119-132
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• 2001

An immunoelectron microscopy employing immunogold labeling method was performed to detect tissue origin of Dl fraction (DIA) among 5 antigenic protein fractions partially purified by DEAE- anion exchange chromatography from water- soluble crude antigen (PIWA) of adult Paragonimus iloktsuenensis. Immune reactions of adult worm tissues with rabbit serum immunoglobulin immunized with crude antigen (PI-Ig) and D1 antigen (D1-Ig), as well as rat serum immunoglobulin infected with P. iloktsuenensis were observed. DlA showed strong antigenicity in the intestinal epithelium of the worms during the early infection period of 2-4 weeks after infection. The vitellaria also showed stronger antigenicity than the other tissue sites in immune reaction of tissues against all immunoglobulins from 4 to 33 weeks after vitelline development. Therefore, it is suggested that DlA was mainly originated from the intestinal epithelial tissues before the development of vitelline gland of the parasites. Immune-reactivity of two immunoglobulins (PI-Ig, Dl-Ig) was significantly different in intestinal epithelial cytoplasmic protrusions (CP) and intestinal epithelial secretory granules (SG). In the experimental group with Dl-Ig, gold particles were labeled significantly in CP than in SG when compared to the PI-Ig group. Thus, the major antigenic materials in Dl antigen having a strong antigenicity in the early infection period was considered to be originated from the intestinal epithelial tissue .

• Parasites, Hosts and Diseases
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• v.51 no.4
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• pp.433-440
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• 2013

Toxocariasis is a worldwide zoonosis caused by larvae of ascarid nematodes of dogs or cats, Toxocara canis or T. cati. Diagnosis of human toxocariasis currently relies on serology that uses T. canis excretory-secretory antigen to detect specific IgG antibodies by ELISA. We investigated the serodiagnostic efficacy of ELISA using crude antigen of T. canis larvae (TCLA). Serum specimens of 64 clinically confirmed toxocariasis, 115 healthy controls, and 119 other tissue-invading helminthiases were screened by ELISA using TCLA. The ELISA using TCLA showed 92.2% (59/64 patient samples) sensitivity and 86.6% (103/119) specificity. Its positive diagnostic predictivity was 78.7% and negative predictivity was 97.8%. No serum of healthy controls reacted but that of anisakiasis (45.5%), gnathostomiasis (19.2%), clonorchiasis (15.8%), sparganosis (11.1%), and cysticercosis (6.3%) cross-reacted. Immunoblot analysis on TCLA recognized antigenic proteins of 28- and 30-kDa bands in their dominant protein quantity and strong blotting reactivity. The present results indicate that the ELISA using our TCLA antigen is acceptable by the sensitivity and specificity for serodiagnosis of human toxocariasis. ELISA with TCLA is recommended to make differential diagnosis for patients with any sign of organ infiltration and eosinophilia.

• Journal of Applied Biological Chemistry
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• v.60 no.1
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• pp.55-59
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• 2017

Although wheat is a common staple food in the world, some people suffer from a variety of wheat allergies. For example, wheat-dependent exercise-induced anaphylaxis is induced in the gastrointestinal tract by wheat proteins. Relatively high molecular weight proteins that are salt-insoluble induce many wheat allergies. In the present study, we investigated the induction of an allergy response using crude wheat proteins, which are relatively low molecular weight, salt-soluble proteins. The crude antigen used in this study was extracted using phosphate buffered saline. When the antigen extracts from various wheat cultivars were orally administered, differentiable degrees of allergy responses were observed as measured by serum IgE and histamine secretion compared to the control. Serum IgE levels increased following administration of three of the wheat extracts. This evidence suggests that a combination of salt-soluble wheat proteins could be antigens for the induction of various allergy responses.

• Biomedical Science Letters
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• v.4 no.1
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• pp.73-76
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• 1998

The authors characterized the antigen proteins and some specific antibodies from Echinostoma hortense. Crude antigen extracted from E. hortense worm was analyzed by SDS-PAGE of the crude antigen showed 46 profiles between 200.2 - 8.2kDa, among which 200.2, 107.9, 86.8, 75, 69.8, 46.8, 43.5, 34.5, 20.9, 13.6, 12.6, 11.7, and 8.2kDa, protein profiles were strong. EITB resolved the specific IgG antibody into 17 profiles between 193 - 13.7kDa, among which 198, 123.4, 100.8, 91.1, 88.1, 62.8, 34.2, 32, 29.9, 18, 15.7, 13.7kDa profiles showed strong immunostain.

• Parasites, Hosts and Diseases
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• v.40 no.4
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• pp.195-198
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• 2002

This study was performed to investigate whethere there is cross-reactivity between Dirofilaria immitis and three intestinal nematodes of dogs. In ELISA, D. immitis- infected dog sera obtained at the 4th molting stage (9-11 weeks) and microfilaremic stage (25-30 weeks) were shown to be highly reactive with crude extract of T. canis. In immunoblotting, some antigenic fractions, 44, 57 88, 100 kDa of crude extract of T. canis, were found to be positive reaction with sera of dogs infected with D. immitis. However, little or no cross-reaction were observed between sera of D. immitis-infected dogs and crude extract antigen of T. vulpis or A. caninum. These result suggest that there are partial cross reaction between sera of D. immitis-infected dogs and the antigen of T. canis.