• Korean Journal of Veterinary Research
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• v.57 no.1
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• pp.31-36
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• 2017

Japanese encephalitis (JE) is an important zoonosis caused by the mosquito-transmitted JE virus (JEV), which is a causative agent of reproductive failure in pregnant sows. Detection of JEV antibodies in swine is performed by hemagglutination inhibition (HI), virus neutralization (VN), and the plaque reduction neutralization test (PRNT). The most stringent PRNT is the 90% endpoint PRNT ($PRNT_{90}$). These conventional assays are difficult to carry out in diagnostic laboratories with insufficient instruments or cell culture systems. An alternative assay that is easily conducted and time efficient is required. In this study, we improved the indirect enzyme-linked immunosorbent assay (I-ELISA) with clarified antigen for the detection of JEV antibodies. The I-ELISA results obtained from 175 swine serum samples were compared with HI, VN, and $PRNT_{90}$ results. The sensitivity of I-ELISA was 91.8%, 95.0%, and 94.7% compared with HI, VN, and $PRNT_{90}$ results, respectively. The specificity of I-ELISA was 92.2%, 94.7%, and 94.7% compared with HI, VN, and $PRNT_{90}$ results, respectively. Moreover, the I-ELISA results were significantly correlated with the HI (r = 0.93), VN (r = 0.95), and $PRNT_{90}$ (r = 0.92) results. These results suggest that the improved I-ELISA is useful for serosurveillance of JEV in swine.

• IMMUNE NETWORK
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• v.21 no.6
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• pp.39.1-39.18
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• 2021

The high virulent severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus that emerged in China at the end of 2019 has generated novel coronavirus disease, coronavirus disease 2019 (COVID-19), causing a pandemic worldwide. Every country has made great efforts to struggle against SARS-CoV-2 infection, including massive vaccination, immunological patients' surveillance, and the utilization of convalescence plasma for COVID-19 therapy. These efforts are associated with the attempts to increase the titers of SARS-CoV-2 neutralizing Abs (nAbs) generated either after infection or vaccination that represent the body's immune status. As there is no standard therapy for COVID-19 yet, virus eradication will mainly depend on these nAbs contents in the body. Therefore, serological nAbs neutralization assays become a requirement for researchers and clinicians to measure nAbs titers. Different platforms have been developed to evaluate nAbs titers utilizing various epitopes sources, including neutralization assays based on the live virus, pseudovirus, and neutralization assays utilizing recombinant SARS-CoV-2 S glycoprotein receptor binding site, receptor-binding domain. As a standard neutralization assay, the plaque reduction neutralization test (PRNT) requires isolation and propagation of live pathogenic SARS-CoV-2 virus conducted in a BSL-3 containment. Hence, other surrogate neutralization assays relevant to the PRNT play important alternatives that offer better safety besides facilitating high throughput analyses. This review discusses the current neutralization assay platforms used to evaluate nAbs, their techniques, advantages, and limitations.

• Biomedical Science Letters
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• v.6 no.2
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• pp.141-149
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• 2000

The etiologic agents of haemorrhagic fever with renal syndrome (HFRS) in Korea are Hantaan and Seoul virus in the genus Hantavirus, family Bunyaviridae. Antibody titers of sera from HFRS patients against Hantaan virus were measured by immunofluorescent antibody technique (IFAT), enzyme-linked immunosorbent assay (ELISA), high density composite particle agglutination (HDPA) and plaque reduction neutralization test (PRNI). PRNT and nested reverse transcriptase polymerase chain reaction (nested RT-PCR) was used for serotypic differentiation of Hantaviruses against Hantaan and Seoul virus. Eight doubtful HFRS patients showed higher fluorescent, IgG ELISA, agglutination and neutralizing antibody titer by IFAT, ELISA IgG, HDPA and PRNT, respectively Five out of them showed high IgM antibody titer by IgM capture ELISA against Hantaan virus, remarkably. Fifteen HFRS patients showed higher fluorescent antibody titer by IFAT. In PRNT, 12 out of them showed high neutralizing antibody titer against HTNV, 2 against SEOV and 1 against both viruses. In nested RT-PCR using serotype specific-primer, 3 out of them showed positive against HTNV and 1 against SEOV.

• International Journal of Oral Biology
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• v.39 no.1
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• pp.35-40
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• 2014

Halitosis is caused by consumption of certain foods or drinks and production of volatile sulfur compounds (VSCs) by periodontopathogens. VSCs-related halitosis is not easily removed using mechanical or chemical therapies such as dental floss, plaque control and mouth rinse. Lactobacillus are known to be probiotics and stimulate immune systems of human. Furthermore, L. casei ATCC 334 and L. rhamnosus GG have an effect on protection of dental caries in vitro studies. The aim of this study was to investigate effect of Lactobacillus on halitosis by Fusobacterium nucleatum- and Porphyromonas gingivalis-producing VSCs and to analyze inhibitory mechanism. The periodontopathogens were cultivated in the presence or the absence Lactobacillus, and the level of VSCs was measured by gas chromatograph. For analysis of inhibitory mechanisms, the susceptibility assay of the spent culture medium of Lactobacillus against F. nucleatum and P. gingivalis was investigated. Also, the spent culture medium of Lactobacillus and periodontopathogens were mixed, and the emission of VSCs from the spent culture medium was measured by gas chromatograph. L. casei and L. rhamnosus significantly reduced production of VSCs. L. casei and L. rhamnosus exhibited strong antibacterial activity against F. nucleatum and P. gingivalis. The spent culture medium of L. casei inhibited to emit gaseous hydrogen sulfide, methyl mercaptan and dimethyl sulfide from the spent culture medium of periodontopathogens. However, the spent medium of L. rhamnosus repressed only dimethyl sulfide. L. casei ATCC 334 may improve halitosis by growth inhibition of periodontopathogens and reduction of VSCs emission.

• Journal of Periodontal and Implant Science
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• v.32 no.3
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• pp.665-683
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• 2002

A novel glucanhydrolase from a mutant of Lipomyces starkeyi KSM 22 has additional amylase activity besides mutanolytic activity and has been suggested as promising anti-plaque agent. It has been shown effective in hydrolysis of mutan, reduction of mutan formation by Streptococcus mutans and removal pre-formed sucrose-dependent adherent microbial film and has been strongly bound to hydroxyapatitie. These in vitro properties of Lipomyces starkeyi KSM 22 glucanhydrolase are desirable for its application as a dental plaque control agent. In human experimental gingivitis　model and 6 month clinical trial, mouthrinsing with Lipomyces　starkeyi KSM 22 dextranase was comparable to 0.12% chlorhexidine mouthwash in inhibition of plaque accumulation and gingival inflammation and local side effect was negligible. This study was aimed to evaluate the cytotoxic effect of Lipomyces starkeyi KSM 22 glucanhydrolase on human gingival fibroblasts. Primary culture of human gingival fibroblasts at the 4th to 6th passages were used. Glucanhydrolase solution was made from lyophilized glucanhydrolase powder from a mutant of Lipomyces stakeyi KSM 22 solved in PBS and added to DMEM medium to the final concentration of 0.5, 1, and 2 unit. Cells were exposed to glucanhydrolase solution or 0.1 % chlorhexidine and the cells cultured in DMEM with 10% FBS and 1% antibiotics as control. After exposure, the morphological change, cell attachment, and cell activity by MTT assay were evaluated in 0.5, 1.5, 3, 6, 24 hours after treatment. The cell proliferation and cell activity was also evaluated at 2 and 7 days after 1 minute exposure, twice a day. The cell morphology was similar between the Lipomyces smkeyi KSM 22 glucanhydrolase groups and control group during the incubation periods, while most fibroblasts remained as round cell regardless of incubation time in the chlorhexidine group. The numbers of the attached cells in the glucanhydrolase groups were comparable to that of control and significantly higher than the chlorhexidine group. The numbers of the proliferated cells in the glucanhydrolase groups at 7 days of incubation were comparable to the control group and higher than the chlorhexidine group. The cell activity in glucanhydrolase groups paralleled with the increased cell number by attachment and proliferation. According to these results, Lipomyces starkeyj KSM 22 glucanhydrolase has little harmful effect on attachment and proliferation of human gingival fibroblasts, in contrast to 0.1% chlorhexidine which was cytotoxic to human gingival fibroblasts. Therefore this glucanhydrolase preparation is considered as a safe and promising agent for new mouthwash formula in the near future.

• Journal of Periodontal and Implant Science
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• v.25 no.3
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• pp.478-486
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• 1995

Periodontal therapy for treatment of periodontitis involves the elimination of bacterial plaque and elimination of the anatomic defects by regenerative procedure. The purpose of this study was to evaluate on the biological effect of magnolia and Ginkgo biloba extract to the antimicrobial, antiinflammatory and cellular activity. Antimicrobial assay was performed with the diffusion method of the extract by measuring of growth inhibitory zone of B. cereus from blood agar plate. Effect of the extract to cellular activity of gingival fibroblast were examined using MTT method and measured the result with optical density on 570nm by ELISA reader. Inhibitory effects of $PGE_2$ production from gingival fibroblast was performed with the addition of $IL-l{\beta}$ and the extract to the well and examined to the product of $PGE_2$ from cell by ELISA reader. In vivo anti-inflammatory effect was performed with injection examined with clinically and histologically for their extent of mecrosis and inflammation. Antimicrobial activity of Magnolia extract showed significantly higher activity than that of control. However, GBE did not showed significant activity to compare with control, and mixture of Magnolia and GBE extract showed significantly higher activity than that of control. The effect of cellular activity to gingival fibroblast showed no significant differences of between control and Magnolia extract. However, GBE showed significantly higher rate of cellular activity to compare with control and even to PDGF-BB, and also showed same degree of cellular activity even though mixed with Magnolia extract. The inhibitory effect of $PGE_2$ production showed significantly reduction of $PGE_2$ production to compare with control, but its inhibitory effect was not much strong to compare with Indomethacin. In vivo, antiinflammatory effect of Magnolia extract to P. gingivalis injection of Hamster buccal check showed significantly reduction of inflammatory cell infiltration and tissue necrosis, but GBE showed no effect on the inhibition of inflammatory process. These results suggested that Magnolia and GBE extract possessed different kind of biological activity and also can be compensated on their activity with each other for elimination of bacterial plaque and anatonical defect.

• Parasites, Hosts and Diseases
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• v.56 no.1
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• pp.61-70
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• 2018

We developed a Rapid Diagnostic Test (RDT) kit for detecting IgG/IgM antibodies against Zika virus (ZIKV) using monoclonal antibodies to the envelope (E) and non-structural protein 1 (NS1) of ZIKV. These proteins were produced using baculovirus expression vector with Sf9 cells. Monoclonal antibodies J2G7 to NS1 and J5E1 to E protein were selected and conjugated with colloidal gold to produce the Zika IgG/IgM RDT kit (Zika RDT). Comparisons with ELISA, plaque reduction neutralization test (PRNT), and PCR were done to investigate the analytical sensitivity of Zika RDT, which resulted in 100% identical results. Sensitivity and specificity of Zika RDT in a field test was determined using positive and negative samples from Brazil and Korea. The diagnostic accuracy of Zika RDT was fairly high; sensitivity and specificity for IgG was 99.0 and 99.3%, respectively, while for IgM it was 96.7 and 98.7%, respectively. Cross reaction with dengue virus was evaluated using anti-Dengue Mixed Titer Performance Panel (PVD201), in which the Zika RDT showed cross-reactions with DENV in 16.7% and 5.6% in IgG and IgM, respectively. Cross reactions were not observed with West Nile, yellow fever, and hepatitis C virus infected sera. Zika RDT kit is very simple to use, rapid to assay, and very sensitive, and highly specific. Therefore, it would serve as a choice of method for point-of-care diagnosis and large scale surveys of ZIKV infection under clinical or field conditions worldwide in endemic areas.

• Archives of Pharmacal Research
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• v.24 no.1
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• pp.74-78
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• 2001

A preparation of water soluble components (EA) was made from carpophores of Elfvingia applanata (Pers.) Karst and its in vitro antiviral activity on vesicular stomatitis virus [(Indiana serotype, VSV(IND)] was investigated by plaque reduction assay. EA exhibited potent antiviral activity on VSV(IND) growth and negligible cytotoxicity on Vero cells, 50% effective concentration ($EC_{50}C$/) of 104$ug\textrm\/ml$ and 50% cytotoxic concentration ($CC_{50}C$) of 3,793$ug\textrm\/ml$, respectively. Selectivity index (Sl $CC_{50}C$/$EC_{50}C$) of EA on Vero cell and VSV(IND) was about 36.5. EA did not display either a direct virucidal effect on V5V(IND) or induction of antiviral substance by Vero cells upon its treatment. Thus, the mode of antiviral activity of EA was studied at steps of viral adsorption onto cell. When both EA and virus were added to cell monolayers, titer of cell-free virus in culture supernatant increased in ca. 30-40% compared with that of control group and titer of cell-associated virus was 60-100% higher than that of control group. These results suggested that antiviral activity of EA on VSV(IND) might be due to the hindrance of viral entry to cells at eITher endocytosis or loss of envelope.

• The Journal of Korean Society of Virology
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• v.30 no.1
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• pp.11-18
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• 2000

Since $Hantavax^{TM}$, formalin inactivated Hantaan virus vaccine (10,240 ELISA units/ml), has been developed in 1990 to prevent against haemorrhagic fever with renal syndrome (HFRS) caused by Hantaan or Seoul virus, it has been commercially available in Korea. Twenty-one healthy people were booster shot once and twice after primary basic vaccination with $Hantavax^{TM}$. Seroconversion rates were measured by immunofluorescent antibody technique (IFAT), enzyme-linked immunosorbent assay (ELISA), high density composite particle agglutination (HDPA), and plaque reduction neutralization test (PRNT). Seroconversion rates of 21 vaccinees at one year after primary basic vaccination were 52.3%, 95.2%, 0.0%, 47.6%, and 28.6%, and 13 vaccinees at one month after 1st booster vaccination were 100%, 100%, 30.7%, 100% and 100% by IFAT, ELISA (IgG, IgM), HDPA and PRNT, respectively. Seroconversion rates declined slightly by twenty months, and they were 84.6%, 92.3%, 0.0%, 84.6% and 69.2% by IFAT, ELISA (IgG, IgM), HDPA and PRNT, respectively. Seroconversion rates of 9 vaccinees at three months after 2nd booster vaccination were 100%, 100%, 0.0%, 100%, and 88.9%, and 16 vaccinees at one year after the 2nd booster vaccination were 87.5%, 93.8%, 0.0%, 87.5% and 81.3% by IFAT, ELISA (IgG, IgM), HDPA and PRNT, respectively. Based on the above result $Hantavax^{TM}$ has proved a vigorous anamnestic response after the 1st and the 2nd booster vaccination and has persisted higher fluorescence, agglutination and neutralizing antibody titers in vaccinees.

• The Korea Journal of Herbology
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• v.30 no.6
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• pp.55-61
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• 2015

Objectives : Yongdamgosam-hwan(YGH) has been used as a traditional medicine from old times for antiinflammatory effects. Streptococcus mutans(S. mutans) is known as a prime bacteria responsible for causing caries by forming a biofilm referred to as dental plaque on the tooth surface. But antimicrobial activity of YGH with dental disease is not sufficiently understood. This study was designed to investigate the effects of YGH ethanol extract on antimicrobial effect against Streptococcus mutans.Methods : The antimicrobial effect of YGH ethanol extract was assessed by the paper disk diffusion method and optical density method to determine minimum inhibition concentration(MIC), also observed by fractional inhibitory concentration index(FICI) and time-kill assay to figure out the synergic effect on the combination of YGH ethanol extract with antibiotics.Results : The YGH ethanol extract 500 μg was 7.5-8.5 mm diameter of clear zone of inhibition against Streptococcus mutans in a concentration-dependent manner and MIC was 250 μg/mL. The administration of the ethanol extract in combination with gentamicin and streptomycin induced a reduction of ≥4-8-fold in all tested bacteria. Furthermore, time-kill study was found that a combination of YGH ethanol extract with oxacillin and streptomycin produced a more rapid decrease in the concentration of bacteria CFU/mL than the YGH ethanol extract or antibiotics alone.Conclusions : As a result, the YGH ethanol extract has good antimicrobial effects. And the results suggest that YGH could be employed as a natural antibacterial agent in dental care products.