• Parasites, Hosts and Diseases
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• v.24 no.2
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• pp.145-158
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• 1986

This study was undertaken to purify cystic fluid (CF) antigen of Taenia solium metacestodes by affinity chromatogaphy using specific monoclonal antibody(McAb) and to characterize the antigenicity of the purified antigen. The hybridoma cell lines, prepared by fusion between mouse plasmacytoma and spleen cells from BALB/c mice immunized with CF, secreted antibodies reacting to various helminthic antigens. Majority of cell lines reacted to CF only but some also reacted to parenchymal antigen of T. solium metacestodes, adult T. saginata, sparganum, hydatid cystic fluid, Paragonimus westermani and Clonorchis sinensis, either in combination with CF, other antigens or independently. Cloned cells derived from monoclonal lines also produced antibodies reacting either to CF only or to other helminthes in combination or independently. These results indicated that CF of T. solium metacestodes contained proteins which possessed antigenic determinants not only specific to CF but also cross reactive with the afore-mentioned helminthes. CF of T. solium metacestodes was purified by affinity chromatography using the McAb which reacted to CF and parenchymal antigens. The affinity-purified antigen (A-Ag) and unbound pool CF (U-Ag) were separated. A-Ag showed 2 protein bands by disc-PAGE whereas CF exhibited 6 bands and U-Ag consisted of all bands CF had. The diagnostic significance of A-Ag was evaluated by ELISA in human neurocysticercosis and other helminthic and neurologic diseases. By A-Ag, the levels of the specific IgG antibody, as shown by absorbance in sera and CSF, were lower than those of CF and U-Ag. Accordingly, the sensitivity was about 70% of CF and U-Ag. However, the nonspecific positive reactions to CF and U-Ag, observed in sparganosis, T. saginata infection and paragonimiasis did not occur when A-Ag was used. These results indicated that the affinity-purified A-Ag had the higher specificity but the lower sensitivity as a diagnostic antigen in cysticercosis, probably because it only detected a single or limited numbers of monospecific antibodies among the diverse polyclonal antibodies produced in the patients with neurocysticercosis.

• Journal of the Institute of Electronics Engineers of Korea SC
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• v.48 no.4
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• pp.102-107
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• 2011

In this paper, a microfluidic array biochip for simultaneously detecting multiple antigen-antibody bindings was designed and implemented. The biochip has the single channel in which microreaction chambers are serially connected, and the antibody-coated microbeads are packed in each microreaction chamber. In addition, the weir structure was fabricated in the microchannel using the gray-scale photolithography in order to trap the microbeads in the microreaction chamber. Three kinds of antibodies were chosen, and the antibodies were immobilized onto the microbeads by the streptavidin-biotin conjugation. In the experiment, as the fluorescence-labeled antigens were injected into the microchannel, the antigen-antibody bindings were completed in 10 minutes. When the solution with multiple antigens was injected into the microchannel, it was observed that the fluorescence intensity increased in only the corresponding microreaction chambers with few non-specific binding. The microfluidic array biochip implemented in this study provides, even with the consumption of tiny amount of sample and fast reaction time to simultaneously detect multiple immunoreactions.

• Parasites, Hosts and Diseases
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• v.34 no.4
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• pp.279-282
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• 1996

The enzyme-linked immunosorbent assay (ELISA)-inhibition test using a Clonorchis sinensis species-specific mouse monoclonal antibody (MAb) , CsHyb 0605-23, showed increased specificity over the conventional ELISA used for serodiagnosis of clonorchiasis. To characterize the corresponding antigen further, the MAb was tested against polysaccharide, protein and glycolipid fractions obtained from a crude extract of C. slnensis adult worms, using chloroform, methanol and phenol extractions. Only the polysaccharide fraction was recognized by the mb among those fractions. Mild oxidation of the antigen with sodium periodate showed decreased reactivity against the MAb. We concluded that the antigen and antigenic determinants recognized by the MAb are carbohydrates.

• Parasites, Hosts and Diseases
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• v.29 no.4
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• pp.339-354
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• 1991

Localization and characterization of the antigenic components of sparganum which induced IgG and IsM antibodies in the host were studied by immunohistochemical techniques and SDS-PAGT and Western blotting. The antigen recognized by IgG antibody of rats or mice which were immunised by infection or injection of crude extracts of metacestodes of Spirometra erinacei, was located in the parenchyme of sparganum, especially at the cortex and around the calcareous corpuscles. The immunoreaction was demonstrated not only in the encysted fibrous wall of host but around the arterioles or venules in the connective tissue of host. The antigen recognized by IgM antibody of rats or mice was also observed in the parenchyme of sparganum and in the connective tissue of host. By 5∼20% gradient SDS-PAGE and EIBT, we detected antigenic components by IgG and 1gG antibodies of the rat or mouse immunized by infection or injection of crude extract of spargana. Twenty-three antigenic bands from crude extracts of spargana were recognized by IgG antibody and 15 components by IgM antibody of immunized rats. Out of the bands recognized by IgG and IgM antibodies, 15 were cross-reacted each other. Twenty components of eBlcretory-secretory proteins from spargana were recognized by IgG, and 5 components by IgM antibody of immunized rats. By IgG and IgM antibodies of immunized mice, 16 components of crude extracts were recognized by IgG antibody and 9 components by IgM antibody. Twenty components of excretory-secretory preparation were recognized by IgG antibody and 5 components by IgM antibody. Thirteen components of crude extracts were cross-reacted by IgG antibody of rats and mice.

• Korean Journal of Animal Reproduction
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• v.15 no.2
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• pp.149-155
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• 1991

본 실험은 H-Y 항원 유전자 크로닝을 위한 기초연구로서 H-Y 항원의 특성을 규명하기 위하여 친화성 크로마토그래피에 의하여 H-Y 항원을 분리·정제하였다. 정소 추출액을 항체가 결합된 column에 결합시킨 뒤 10% acetic acid로 용출시켰다. 용출된 분획을 모아 농축한 후 HPLC와 SDS-PAGE를 실시하여 H-Y 항원의 분자량은 약 6,7000달톤 임을 알 수 있었으며 isoelectric focusing에 의하여 등전점(pI)은 5.0인 것으로 측정되었다. H-Y 항원에 대한 단일클론항체와 표지항원으로는 H-Y 항원-ABEI(aminobutylethyl isoluminol)를 사용하여 H-Y 항원 정량을 위한 화학발광면역분석법을 개발하였다. 항원항체 반응후 빛의 측정은 NaOH 존재하에서 microperoxidase/H2O2를 이용한 산화반응으로 실시하여 10초간 측정한 빛의 양을 적분하였다. H-Y 항원의 농도와 빛의 양과는 역비례하였으며 감도는 11.8ng/tube 정도이었다.

• The Korean Journal of Zoology
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• v.32 no.4
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• pp.429-440
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• 1989

Spirometru erinocei의 유충인 sparganum에서 추출한 조항원 단백질을 항원으로 하여 sparganosis, cysticercosis, hydatidosis환자의 IgG 항체와 정상인의 IgG 항체를 혈청반응시켜 ELISA와 EITB에 의해 교차반응을 일으키는 비 특이항원 성분과 종특이항원 성분을 추구하였다. 1. sparganum에서 0.01 M PBS (PH 7.4)로 추출한 조항원 단백질을 SDS-PAGE로 전개하여 290 Kd에서 23 Kd범위의 분자량을 가진 25개의 단백질이 분획되었다. 2. sparganum의 조항원을 항원으로하여 ELISA로 sparganosis, cysticercosis, hydatidosis 환자의 190 항체와의 혈청 반응치는 sparganosis환자에게서는 0.44 $\pm$ 0.07에서 1.90 $\pm$ 0.03으로 negative control normal sera의 0.D (0.15 $\pm$ 0.03)를 기준으로 하였을 때 모두 양성이며 민감도(sensitivity)가 100 %이었으며 cysticercosis, hydatidosis환자혈청에서 양성반응이 나타났으며 교차반응도 있었다. 3. EITB에서는 spargancsis환자의 IgG항체에 의해 16개의 항원성분이 인지되었으며 이 중 6개의 항원성분이 정상인의 혈청에서도 인지되어 교차반응을 일으키는 항원성분이었으며 cysticercosis환자혈청에서 인지된 4개의 항원성분 중 2개의 항원성분이 sparganosis 환자혈청에서 인지된 것과 같았으며 hydatidosis환자의 190항체에 의해 인지된 19개의 항원성분 중 12개의 항원성분이 sparganosis환자혈청에서 인지된 항원 성분과 같았다. 4. 290 Kd, 200 Kd, 28 Kd의 항원성분은 sparganosis환자의 196항체에서만 인지되었고 228 Kd, 152 Kd, 66 Kd항원성분은 hydatidosis환자의 19G항체에서만 인지되었으며 66 Kd항원성분은 sparganosis, cysticercosis, hydatidosis, 정상인의 혈청에서 모두 인지되었다. We studied the serological reaction between the specific and nonspecific antigenic components from metacestode (plerocercoid) of spiromeko erimacei and IgG antibodies in sparganosis, cysticercosis. hydatidosis patients and normal human sera by ELISA and EITB. We prepared the crude extracts of sparganlim from snake, Matrix tigrina laterolis and used as antigenic components. By SDS-PAGE, we detected a total 25 peptide bands (fractions) with 290 Kd to 23 Kd molecular weight, and 8 bands of these detected bands developed strongly by silver stain. In serological test, ELISA, we recognized the cross-reaction of antigenic components reacting with IgG antibodies in heterogenous sera, cysticercosis and hydatidosis patients sera. The crude antigenic components of sparganum showed the high sensitivity in sparganosis, hydatidosis patients sera, but showed lower sensitivity in cysticercosis patients sera than the sparasanosis, hydatidosis patients sera. Sixteen antigenic components of these 25 separated bands were recognized by antibodies In sparsanosis patients sera,8 antigenic components in normal human sera, 4 antigenic components in cysticercosis patients sera and 19 antigenic compoenents in hydatidosis patients sera. The crude antigenic compnenets with 290 Kd, 200 Kd, 125 Kd and 28 Kd molecular weight was only recognized in sparganosis patients sera, but 64 Kd antigenic component was nonspecific antigenic components which were also cross-reacted with sparganosis, hydatidosis, cysticercosis patients sera and normal human sera.

• Proceedings of the Korean Vacuum Society Conference
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• 2012.02a
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• pp.558-558
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• 2012

최근의 원자간력현미경(AFM)은 soft한 생체물질을 비파괴적 방법 및 나노크기의 분해능으로 여러 구조적, 물리적 특성 측정이 가능하여 bio분야에 다양이 활용되고 있다. 본 연구에서는 AFM을 이용하여 줄기세포인 BM MSC(bone marrow mesenchymal stem cell)가 신경세포로 분화 여부를 측정하는 방법을 보고하고자 한다. 신경세포의 신호전달은 시냅스에서 신경전달물질을 매개로 하여 이루어지는데, 신경전달물질 중에 D-Glutamic acid는 시냅스후세포에서 흥분성 전위 크기를 증가시킨 상태를 장기간 유지시켜주는 물질로, 특정물질인 Glutamate와 항원-항체 결합을 한다. 본 연구에서는 이 두 물질간의 항원-항체 반응을 활용하여 줄기세포의 신경세포로 분화 여부를 AFM으로 측정하였다. 먼저, 수용성 시료인 두 물질을 증류수에 용해시켜 Mica 기판에 그 용액을 떨어뜨려 자연건조로 시료를 준비한 후, AFM으로 형태 및 크기를 측정하였다. D-Glutamic acid와 Glutamate는 구형 입자 형태를 보였으며, Glutamate의 너비는 ~100 nm이고, D-Glutamic acid는 ~50 nm였다. 두 물질이 든 용액을 섞었을 때, 항원-항체 반응에 의해 다른 크기의 두 구형입자가 붙어 있는 형태가 관찰되었다. 이 반응을 활용하여, 신경세포에서 분비되는 신경전달물질인 D-Glutamic acid를 선별하였다. DMEM 배지에 신경암세포주인 SH-SY5Y 를 접종한 후 $37.6^{\circ}C$의 incubator에서 24시간 배양하고, 화학적 자극(60~70 mM의 KCl 용액을 주입함)을 주어 신경전달물질 분비를 유도하였다. 그 배지에 항체 Glutamate 를 주입하여 자연건조 시킨 후 항원-항체 결합특성을 AFM으로 측정하여, 항원-항체 결합된 이미지와 동일함을 확인하였다. 결과적으로 AFM을 이용한 신경전달물질의 항원-항체 결합여부 측정을 통해, BM MSC 줄기세포의 신경세포로 분화를 판단할 수 있으며, 이 방법은 줄기세포의 특정 세포로의 분화 여부 판단에 활용될 것으로 기대된다.

• Parasites, Hosts and Diseases
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• v.30 no.1
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• pp.1-14
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• 1992

In order to observe the antigenic localization in the tissues of Paragonimus westermani of deve- lopmental stages, immunogoldlabeling method was applied using serum of the cats which were infected with isolated metacercariae from Cambaroides similis. The sectioned worm tissues from orch developmental stage were embedded in Lowicryl HM20 medium, stained with infected semi IgG and protein A gold complex(particle size: 12 nm) and observed by electron microscopy. In the young adult worm tissue of 4 weeks after infection with metacercariae, the gold particles were specifically concentrated on the tegumental syncytium and cytoplasm of the tegumental cells as well as the secretory granules in the parenchymal tissue. The antigenic materials in the adult worm tissue were specifically concentrated on the secretory granules in the parenchymal tissue, the cytoplasm between granules in the vitelline gland and the epithelial lamella in the lumen of the caecum.

• Journal of Sericultural and Entomological Science
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• v.15 no.2
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• pp.1-7
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• 1973

It was found that the diapause in the silkworm egg is induced by the action of the diapause hormone secreted from the suboesophageal ganglion, and "esterase A" affects protein metabolism in oocyte and egg. In this connection, some changes in protein metabolism of silkworm egg according to embryonic developments could give some information on the diapause, using Ouchterlony Test. Antigenicity of the protein of silkworm egg was detected through antigen-antibody interaction among the extracts of rabbit blood. Furthermore, existence of the specific antigen was also detected according to embryonic development, using the adsorption test. The results were obtained as follows: 1 Detection of antigenicity The antigenicity of silkworm egg was ascertained by inoculating it into a rabbit, but positive results were shown in most of the silkworm eggs tested, whereas the antibody specific to a certain antigen was not detected. 2. Detection of the common antigen It was demonstrated that most of the antigen could incite the common antibodies, but the specific antibody formation was not detected in a few antigens, even though the nonspecific antibody formation was displayed. 3. Detection of the specific antigen It is suggested that there are the specific antigens detectable in each treated eggs by the adsorption test.

• Parasites, Hosts and Diseases
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• v.29 no.3
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• pp.293-310
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• 1991

Clonorchis sinensis is a common parasite of man in Korea. Researches on the specific antigens of C. sinensis would be valuable not only because those elucidate the molecular characteristics of this fluke but also because it is applicable to immunodiagnosis. Although many monoclonal antibodies have been used in the field of parasite immunology, few articles on monoclonal antibodies against C. sinensis have been published so far. The aim of this study was to analyse C. sinensis antigens recognized by monoclonal antibodies, and to set up ELISA-inhibition test using C. sinensis specific monoclonal antibodies for improved specificity of immunodiagnostic tests. By fusion between spleen cells of the mice immunized with C. sinensis water-soluble crude adult worm antigens and plasmacytoma cells of mouse origin, 29 hybridoma clones secreting anti-C. sinensis monoclonal antibodies were made, and 8 clones among those were found specific. After cell cloning, isotypes of 6 selected specific monoclonal anti- bodies were determined to be IgGl, IgG2b and IgA. Four exposed antigenic determinants of natural infection were recognized by different specific monoclonal antibodies. By enzyme-immunoelectrotransfer blot, 10 KD, 34 KD antigenic determinants were found to be reacted with CsHyb 0714-20, CsHyb 0605-10 monoclonal antibodies, respectively, The antigenic determinant recognized by CsHyb 0714-20 monoclonal antibody was revealed to be located at the surface and parenchyme of a parasite by indirect immunoauorescent antibody technique, and those reacted with CsHyb 0605-10, CsHyb 0714-25 monoclonal antibodies were found at the parenchyme and intestine. The antigenic determinant reacted with CsHyb 0605-23 monoclonal antibody was found mainly around the uterine eggs. Four antigenic determinants recognized by specific monoclonal antibodies were all found to be present in the early eluted fractions of C. sinensis antigens separated by Sephadex G-200 gel filtration. By conventional ELISA, 75% of clonorchiasis cases were found positive, but 7.1% of normal controls and 37.5% of paragonimiasis cases showed false positives. However, by ELISA-inhibition test using C. sinensis specific monoclonal antibody (CsHyb 0605-23), 77.1% of clonorchiasis cases were found positive, and there were no false positives in normal controls or paragonimiasis cases, indicating 100% specificity. The ELISA- inhibition test using monoclonal antibodies was found to have same sensitivity and definitely high specificity in comparison with conventional ELISA for serodiagnosis of human clonorchiasis.