• Journal of the Korea Academia-Industrial cooperation Society
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• v.18 no.1
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• pp.295-301
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• 2017

Botulinum neurotoxin (BoNT/A) is a neurotoxin that selectively attacks the peripheral cholinergic nerve endings. It is produced by Gram -positive, endospore-forming strict anaerobic bacteria, Clostridium botulinum. Since BoNT/A could be a biothreat agent, as well as a contaminator of food and water supplies, the development of sensitive assays for toxin detection and potent antitoxin for the treatment of intoxication is necessary. In this study, for the purpose of producing monoclonal antibodies (mAbs) that are capable of neutralizing Botulinum neurotoxin type A (BoNT/A), scFv (single-chain variable domain fragment) libraries from the rabbit antisera against BoNT/A was fused to a human IgG. The resulting recombinant scFvIgG antibody protein was expressed in stable cell lines and was purified using a protein A affinity chromatography. The efficacy of scFvIgG mAb was confirmed by ELISA and was evaluated for the neutralization of BoNT/A in vivo. Such an in vivo toxin neutralization assay was performed using mice. Although scFvIgG antibody proteins (10 ug) failed to fully protect the mice challenged with BoNT/A (100,000 $LD_{50}$), it significantly prolonged the survival time. These results suggest that scFvIgG mAb may be capable of neutralizing BoNT/A single-chain variable domain fragment.

• Journal of Korean Neurosurgical Society
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• v.63 no.2
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• pp.178-187
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• 2020

Objective : The extensive vasa vasorum network functions as a conduit for the entry of inflammatory cells or factors that promote the progression of angiogenesis and plaque formation. Therefore, we investigated the correlation between the carotid vasa vasorum activities and carotid plaque vulnerability using indocyanine green video angiography (ICG-VA) during carotid endarterectomy (CEA). Methods : Sixty-nine patients who underwent CEA were enrolled prospectively from September 2015 to December 2017. During CEA, a bolus of ICG was injected intravenously before and after resecting the atheroma. Additionally, we performed immunohistochemistry using CD68 (a surface marker of macrophages), CD117 (a surface marker of mast cells), and CD4 and CD8 (surface markers of T-cells) antibodies to analyze the resected plaque specimens. Results : The density of active vasa vasorum was observed in all patients using ICG-VA. The vasa vasorum externa (VVE) and interna (VVI) were seen in 11 (16%) and 57 patients (82.6%), respectively. Macroscopically, the VVE-type patterns were strongly associated with preoperative angiographic instability (81.8%, p=0.005) and carotid plaque vulnerability (90.9%, p=0.017). In contrast, the VVI-type patterns were weakly associated with angiographic instability (31.6%) and plaque vulnerability (49.1%). CD68-stained macrophages and CD117-stained mast cells were observed more frequently in unstable plaques than in stable plaques (p<0.0001, p=0.002, respectively). Conclusion : The early appearance of VVE, along with the presence of many microvessel channels that provided nutrients to the developing and expanding atheroma during ICG-VA, was strongly associated with unstable carotid plaques. The degree of infiltration of macrophages and mast cells is possibly related to the formation of unstable plaques.

• Journal of the korean academy of Pediatric Dentistry
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• v.30 no.1
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• pp.61-68
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• 2003

Streptococcus mutans is known to be a major causative organism of human dental caries. The development of a vaccine against dental caries involves identification of appropriate antigens of mutans streptococci against which protective immune responses can be mounted, and the selection of a method of immunization that will generate sustained levels of protective antibodies. Antigens receiving most attention include streptococcal surface proteins that are involved in attachment to tooth surfaces and glucosyltransferases (GTF) that synthesize adhesive glucans from sucrose. The induction of antibody responses to orally administered antigens is often difficult due to digestive destruction of antigens and immune tolerance. Here we report the induction of antibody responses to an anti-caries vaccine containing retinoic acid (RA). Subcutaneous immunization with formalin-fixed bacteria or GTF supplemented with RA induced higher serum IgM and IgA responses to GTF compaired to oral adminstration. Antisera induced by Ingbritt strain showed partial cross-reaction with LM-7 strain, but not with OMZ175. These results suggest that subcutaneous immunization with GTF combined with an immunomodulator, RA, may be applied to anti-caries vaccine.

• The Journal of Korean Society of Virology
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• v.26 no.1
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• pp.9-22
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• 1996

Respiratory Syncytial virus (RSV) is an important cause of acute lower respiratory tract infections in human, with infants and young children being particularly susceptible. In the temperate zones, sharp annual outbreaks of RSV occur during the colder months, in both the northern and the southern hemisphere. RSV is unusual in that it can repeatedly reinfect individuals throughout life and infect babies in the presence of maternal antibody. RSV isolates can be divided into two subgroups, A and B, on the basis of their reactions with monoclonal antibodies, and the two subgroups are also distinct at the nucleotide sequence level. The specific diagnosis of RSV infection was best made by isolation of virus in tissue culture, identification of viral antigen, or by specific serologic procedures. Recently, rapid detection of RSV and analysis of RSV strain variation became possible by development of methods of reverse transcription and polymerase chain reaction amplification. In this study, to determine the genetic diversity of RSV found in Korea, 173 bp and 164 bp spanning selected regions of the RSV F and SH genes were enzymatically amplified and sequenced, respectively. Eight for F gene and three for SH gene were detected in 66 nasopharyngeal swap samples tested. Two major antigenic subgroups, A and B were confirmed from Korean samples (seven for subgroup A and one for subgroup B). At the nucleotide level of the F gene region, Korean subgroup A strains showed 95-99% homologies compared to the prototype A2 strain of subgroup A and 93-100% homologies among Korean subgroup A themselves. For the SH gene region, Korean subgroup A strain showed 97.5% homology compared to the prototype A2 strain of subgroup A, and Korean subgroup B strain showed 97% homology compared to the prototype 18537 strain of subgroup B. Most of base changes were transition and occured in codon position 3, which resulted in amino acid conservation. Using the maximum parsimony method, phylogenetic analysis indicated that Korean RSV strains formed a group with other RSV strains isolated from the United States, Canada, the Great Britain and Australia.

• Asian Pacific Journal of Cancer Prevention
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• v.14 no.10
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• pp.5973-5981
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• 2013

At present, the most common cause of cancer-related death in women is breast cancer. In a large proportion of breast cancers, there is the overexpression of human epidermal growth factor receptor 2 (HER2). This receptor is a 185 KDa growth factor glycoprotein, also known as the first tumor-associated antigen for different types of breast cancers. Moreover, HER2 is an appropriate cell-surface specific antigen for passive immunotherapy, which relies on the repeated application of monoclonal antibodies that are transferred to the patient. However, vaccination is preferable because it would stimulate a patient's own immune system to actively respond to a disease. In the current study, several bioinformatics tools were used for designing synthetic peptide vaccines. PEPOP was used to predict peptides from HER2 ECD subdomain III in the form of discontinuous-continuous B-cell epitopes. Then, T-cell epitope prediction web servers MHCPred, SYFPEITHI, HLA peptide motif search, Propred, and SVMHC were used to identify class-I and II MHC peptides. In this way, PEPOP selected 12 discontinuous peptides from the 3D structure of the HER2 ECD subdomain III. Furthermore, T-cell epitope prediction analyses identified four peptides containing the segments 77 (384-391) and 99 (495-503) for both B and T-cell epitopes. This work is the only study to our knowledge focusing on design of in silico potential novel cancer peptide vaccines of the HER2 ECD subdomain III that contain epitopes for both B and T-cells. These findings based on bioinformatics analyses may be used in vaccine design and cancer therapy; saving time and minimizing the number of tests needed to select the best possible epitopes.

• BMB Reports
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• v.36 no.6
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• pp.545-551
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• 2003

A cDNA of bovine brain glutamate dehydrogenase (GDH) was isolated from a cDNA library by recombinant PCR. The isolated cDNA has an open-reading frame of 1677 nucleotides, which codes for 559 amino acids. The expression of the recombinant bovine brain GDH enzyme was achieved in E. coli. BL21 (DE3) by using the pET-15b expression vector containing a T7 promoter. The recombinant GDH protein was also purified and characterized. The amino acid sequence was found 90% homologous to the human GDH. The molecular mass of the expressed GDH enzyme was estimated as 50 kDa by SDS-PAGE and Western blot using monoclonal antibodies against bovine brain GDH. The kinetic parameters of the expressed recombinant GDH enzymes were quite similar to those of the purified bovine brain GDH. The $K_m$ and $V_{max}$ values for $NAD^+$ were 0.1 mM and $1.08\;{\mu}mol/min/mg$, respectively. The catalytic activities of the recombinant GDH enzymes were inhibited by ATP in a concentration-dependent manner over the range of 10 - $100\;{\mu}M$, whereas, ADP increased the enzyme activity up to 2.3-fold. These results indicate that the recombinant-expressed bovine brain GDH that is produced has biochemical properties that are very similar to those of the purified GDH enzyme.

• The Journal of Korean Society of Virology
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• v.30 no.2
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• pp.101-112
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• 2000

Rotaviruses are the most common cause of severe vomiting and diarrhea in children worldwide and classified as a genus in the family Reoviridae. Rotavirus has eleven segmented dsRNAs and the virion consists of three shells. Outer capsid VP7 and VP4 induce neutralizing antibodies and are classified into G types (glycoprotein VP7) and P types (protease-sensitive VP4). Characterization of VP7 gene of Korean isolates of human rotavirus was performed using multiplex PCR and nucleotide sequence analysis. After RT-PCR amplification of full length (1,062 bp) of VP7 genes, the amplified PCR products were G typed by multiplex PCR and the nucleotide sequences were compared with those of reference rotavirus from GenBank. The G type analysis revealed that 25% (2/8) belong to G1, whereas 37.5% (3/8) benong to G2 and G4, respectively. The Korean isolates within the same serotypes showed high homology of nucleotide sequences and could be discriminated from foreign isolates exception with two strains (CAU009 and CAU022). But Korean isolates CAU009 and CAU022 were close related into japanease isolates 417 (99.2%) and indian isolates (97.6%) than Korean isolatese. Our results showed that these two strains were supposed to be originated from abroad. As a results, The G typing and nucleotide sequence analysis of VP7 gene of rotavirus isolated from infants in Korea could be used for identification, serotying and determination of novel or unusual strains of rotaviruses.

• Parasites, Hosts and Diseases
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• v.50 no.4
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• pp.287-293
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• 2012

Knowledge of the prevalence of human Toxoplasma gondii infection is required in the Republic of Korea. In this study, we surveyed the seroprevalence of T. gondii infection and analyzed the risk factors associated with seropositivity among residents in 2 administrative districts; Seoul and the island of Jeju-do, which have contrasting epidemiologic characteristics. Sera and blood collected from 2,150 residents (1,114 in Seoul and 1,036 in Jeju-do) were checked for IgG antibody titers using ELISA and for the T. gondii B1 gene using PCR. In addition, participants completed a questionnaire that solicited information on gender, age, occupation, eating habits, history of contact with animals, and travel abroad. The T. gondii B1 gene was not detected in all residents examined. However, ELISA showed 8.0% (89 of 1,114 sera) positive for IgG antibodies against T. gondii in Seoul and 11.3% (117 of 1,036 sera) in Jeju-do. In both districts, the positive rates were higher in males than in females, and those 40-79 years of age showed higher rates than other ages. In Seoul, residents older than 70 years of age showed the highest positive rate, 14.9%, whereas in Jeju-do the highest prevalence, 15.6%, was in those in their sixties. The higher seropositive rate in Jeju-do than in Seoul may be related to eating habits and occupations. The present results and a review of related literature are indicative of an increased seroprevalence of T. gondii in Korea in recent years.

• Parasites, Hosts and Diseases
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• v.51 no.3
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• pp.363-367
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• 2013

The prevalence of Toxoplasma gondii infection in birds has epidemiological significance because birds are indeed considered as a good indicator of environmental contamination by T. gondii oocysts. In this study, the prevalence of T. gondii in 313 house sparrows in Lanzhou, northwestern China was assayed by the modified agglutination test (MAT). Antibodies to T. gondii were positive in 39 (12.46%) of 313 samples (MAT titer ${\geq}$ 1:5). Tissues of heart, brain, and lung from the 39 seropositive house sparrows were tested for T. gondii DNA, 11 of which were found to be positive for the T. gondii B1 gene by PCR amplification. These positive DNA samples were typed at 9 genetic markers, including 8 nuclear loci, i.e., SAG1, 5'- and 3'-SAG2, alternative SAG2, SAG3, GRA6, L358, PK1, c22-8 and an apicoplast locus Apico. Of them, 4 isolates were genotyped with complete data for all loci, and 2 genotypes (Type II variants; ToxoDB #3 and a new genotype) were identified. These results showed that there is a potential risk for human infection with T. gondii in this region. To our knowledge, this is the first report of T. gondii seroprevalence in house sparrows in China.

• Korean Journal of Veterinary Research
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• v.39 no.2
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• pp.247-256
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• 1999

A disease of young Holstein calves characterized by recurrent pneumonia, ulcerative and granulomatous stomatitis, enteritis with bacterial overgrowth, periodontitis, delayed wound healing, persistent neutrophilia and death at an early age had been originally described in 1983 and again in 1987. Most of these calves had stunted growth and a persistent, progressive neutrophilia (often exceeding 100,000/ml). By investigation of pedigrees, all of the affected calves have now been traced to a common sire and confirmed by polymerase chain reaction (PCR) diagnostic DNA testing to be homozygous carriers of a defective allele for bovine CD18. Neutrophils from these calves have several functional deficits and, most importantly, fail to adhere in a ${\beta}_2$-integrin dependent manner. The ${\beta}_2$-integrins represent a family of glycoproteins which participate in various leukocyte adhesion reactions during host defense. The presence or absence of ${\beta}_2$-integrin molecules can be demonstrated on the surface of neutrophils, monocytes and lymphocytes from normal or affected calves using specific monoclonal antibodies and flow cytometry, or by colloidal gold immunolabeling and scanning electron microscopy in backscatter mode. Deficiency of the ${\beta}_2$-integrins on all leukocyte types in Holstein calves is analogous to leukocyte adhesion deficiency (LAD) seen in humans. Neutrophils in bovine (BLAD) and human LAD patients are unable to adhere to the endothelial lining of the cardiovascular system thus interrupting egression of neutrophils into infected tissues. Other leukocytes, while still deficient in expression of the ${\beta}_2$-integrins, are still able to efficiently egress from the blood stream due to interactions of other adhesion molecules that are not as highly expressed on neutrophils. Both BLAD cattle and LAD children (who do not receive bone marrow transplants) often die at an early age as a result of the failure of neutrophils to extravasate into infected tissues. In 1991, Shuster, et $al^{27}$, identified two point mutations within the alleles encoding bovine CD18 in a Holstein calf afflicted with leukocyte adhesion deficiency. One mutation causes an aspartic acid to glycine substitution at amino acid 128 (D128G) in an extracellular region of this adhesion glycoprotein that is highly conserved (> 95% identity) between humans, cattle and mice. The other mutation is silent. Numerous calves with clinical symptoms of leukocyte adhesion deficiency have since been tested and all have been found homozygous for the D128G allele. In addition, calves homozygous far the D128G allele have been identified during widespread DNA testing in the United States. All cattle with the mutant allele are related to one bull, who through artificial insemination (A.I.), sired many calves in the 1950's and 1960's. The carrier frequency of the D128G CD18 allele among U.S. Holstein cattle had reached approximately 15% among active A.I. bulls and 8% among cows. By 1993, the organization of the dairy industry and the diagnostic test developed to genotype cattle, enabled virtually complete eradication of bovine leukocyte adhesion deficiency among current and future A.I. bulls.