• Korean Journal of Veterinary Service
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• v.42 no.1
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• pp.9-15
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• 2019

Transmissible gastroenteritis (TGE) is a disease confined to pigs of all ages, and can be a significant cause of economic loss in breeding herds, primarily because of the very high piglet mortality. The causative agent is a coronavirus, an enveloped positive strand RNA virus and closely related but non-enteropathogenic porcine respiratory coronavirus (PRCV). Although the TGEV has declined with its innocent relative, PRCV, further genome changes could not be excluded. Therefore, the herd-level immunity against this virus is important for the prevention of disease and should be carefully monitored. The aim of this study is to develop monoclonal antibody capture enzyme-linked immunosorbent assay (MAC-ELISA) which can rapidly and accurately determine a large numbers of serum samples for surveillance purpose, and to compare the ELISA with a TGEV-specific serum neutralization test. The MAC-ELISA was sufficiently achieved, and the comparison with the virus-specific serum neutralization assays for 713 sera from pig farms showed a high correlation ($r^2=0.812$, P<0.001). The specificity and sensitivity of MAC-ELISA for the serum neutralization test 91.9% and 91.6%, respectively, which means that the antibody detected by the MAC-ELISA could be said to be protective antibodies. In conclusion, the developed MAC-ELISA would be very helpful in evaluating protective antibodies against TGEV.

• Journal of Pharmacopuncture
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• v.7 no.3
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• pp.5-25
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• 2004

Objectives : To observe changes in the serum proteins before and after intravenous injection of wild ginseng herbal acupuncture. Methods : Blood was collected before and after the administration of wild ginseng herbal acupuncture and only the serum was centrifuged. Then differences in the spots on the scanned image after running 2-Dimensionl electrophoresis were located and conducted mass analysis and protein identification. Results : Following results were obtained from the comparative analysis of serum proteins before and after the administration of wild ginseng herbal acupuncture. 1. 28 spots were identified before and after the administration. 2. In confirming manifestation degree, spots with more than two-times increase were 204, 803, 1505, 2205, 3105, 7104, 9001 spots, with more than one-time increase were 1101, 1302, 2013, 3009, 3010, 4002, 4009, 6706, 7103, 8006, 8101, and spots with decrease were 205, 801, 3205, 5202, 6105. 3. After conducting protein identification, proteins 205, 804, 1302, 4009, 6105, 6106 are unidentified yet, and 1101 is unnamed protein. Protein 204 is identified as complement receptor CR2-C3d, 801 as YAP1 protein, 803 as antitrypsin polymer, 1505 as PRO0684, 2013 and 3010 as proapolipoprotein, 2205 as USP48, 2403 as vitamin D binding protein, 3009 as complement component 4A preprotein, 3105 as immunoglobulin lambda chain, 3205 as transthyretin, 4002 as Ras-related protein Ral-A, 4204 as beta actin, 5202 and 7104 as apolipoprotein L1, 6704 as alpha 2 macroglobulin precursor, 7103 as complement component 3 precursor, 8006 as testis-specific protein Y, 8101 as Transferrin, 9001 as(Alpha-Oxy, Beta-(C112g)deoxy) T-State Human Hemoglobin, and 9003 as human hemoglobin. 4. Immune protein CR2-C3d, which acts against microbes and pathogenic organisms, and Antitrypsin(803), which is secreted with inflammatory response in the lungs, were increased by more than 200% after the administration of herbal acupuncture. 5. Immunoglobulin lambda chain(3105), Alpha-Oxy, Beta-(C112g)deoxy T-State Human Hemoglobin(9001), and human hemoglobin(9003) were increased by more than two-times after the administration of herbal acupuncture. 6. Proapolipoprotein(2013, 3010) and apolipoprotein(7104), key components of the HDL-cholesterol which plays an important role in preventing arteriosclerosis, were increased after the administration of herbal acupuncture. 7. Vitamin D binding protein(DBP, 2403), protecting the lung at the time of inflammatory response, was increased after the administration of herbal acupuncture. 8. Transthyretin(TTR, 3205), which is the main protein causing familial aimyloid polyneuropathy(FAP), was decreased after the administration of herbal acupuncture. 9. Ras-related protein Ral-A(4002) that controls phospholipid metabolism, cytoskeletal formation, and membrane traffic, was increased after the administration of herbal acupuncture. 10. Testis-specific protein Y(8006), which takes part in determination of the gender, was increased by more than two-times after the administration of herbal acupuncture. 11. Transferrin(8101), T-State Human Hemoblobin(9001), and Human Hemoblobin(9003) which balances the iron level in the body, were increased after the administration of herbal acupuncture. Conousion : Above results support the notion that intravenous injection of cultivated wild ginseng herbal acupuncture induce changes in serum proteins and this research can be a pioneer work in finding biomarkers.

• Korean Journal of Veterinary Research
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• v.32 no.1
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• pp.83-90
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• 1992

Monoclonal antibodies against structural proteins of bovine viral diarrhea virus(BVDV) were derived by classical hybridoma techniques. These antibodies were characterized by serum neutralization, immunoblotting and immunoprecipitation. The neutralizing monoclonal antibody reacted with the 56kd to 54kd(M.W.) viral protein in western blotting and immunoprecipitation analysis. Although there was no neutralizing activity, another monoclanal antibody reacted with the 45kd protein by immunoprecipitation and with both the 45kd and 36kd proteins in immunoblotting analysis. respectively. Densitometer scanning of purified BVDV and the immunopreipitation of whole virus particles with neutralizing monoclonal antibody revealed the presence of more than twelve viral polypeptides. Although no possible precursor form of protein was identified with the neutralizing monoclonal antibody. the presence of intact virion was detected in the infected cell supernatant immediatelty after pulse labeling, indicating rapid translational processing as well as packaging of the virus. The partial peptide mapping of 45kd and 36kd proteins with Staphylococcus aureus V 8 protease showed that these two proteins are related.

• Korean Journal of Food Science and Technology
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• v.29 no.5
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• pp.1067-1070
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• 1997

Imunoglobulins from bovine plasma proteins were isolated by IMAC which $Cu^{2+}$ was chelated on a chelating sepharose fast flow gel. Most plasma proteins were eluted by 1st (0.01 M $Na_2HPO_4$, 0.5 M NaCl, pH 4.0) and 2nd elution buffers (0.01 M imidazol). According to the reverse phase HPLC analysis, it was found that proteins which were eluted by 1st elution buffer were mainly composed of serum albumin, while most IgG and transferrin were eluted by 2nd elution buffer. When protein fractions obtained by 2nd elution buffer was applied to ultra filtration system (molecular weight cut off: 100 kD), IgG was further purified. These results indicate that IMAC is an excellent tool for isolating imunoglobulins from plasma proteins.

• Journal of Animal Science and Technology
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• v.47 no.1
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• pp.19-28
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• 2005

The objectives of the current study were to develop polyclonal antibodies in sheep against adipocyte plasma membrane(APM) proteins isolated from swine, to investigate tissue specificity, and to determine cytotoxic effects of antiserum on swine adipocytes. Plasma membrane proteins from adipocyte, brain, heart, kidney, liver, and spleen were isolated using a 32% sucrose gradient. Adult male sheep was immunized three times at three week interval with the purified swine APM proteins. Antiserum was taken from immunized sheep at 10, 12, and 14 days after the third immunization. Antiserum expressed strong reactivity with APM proteins determined by enzyme-linked immunosorbent assay(ELISA), and the reactivity could be detected at dilutions in excess of 1 : 81,000. Antiserum showed very low binding affinity with proteins isolated from brain, heart, kidney, liver, or spleen. Tissue specificity of the antiserum was reconfirmed by Western immunoblotting using anti-sheep immunoglobulin G•alkalinephosphatase conjugate as a secondary antibody. The reactivity of antiserum to the external surface of fixed swine adipocytes was confmned by an immunohistochemical technique using anti-sheep immunoglobulin G-FITC. Confluent swine adipocytes in culture were lysed by antiserum treatment and cytosolie lactate dehydrogenase(LDH) was released as a dose-dependent patterns while adipocytes treated with normal sheep serum maintained their integrity and expressed low level of LDH. These results implicate that fat contents in the pigs can be reduced by immunological methods.

• Microbiology and Biotechnology Letters
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• v.41 no.2
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• pp.242-248
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• 2013

Endotoxins are part of the outer membrane of the cell wall of gram-negative bacteria and are continuously released during bacterial growth. Endotoxins typically induce severe sepsis and septic shock, which cause more than 50% of mortalities. Endotoxins are easily measured in the serum by the limulus amebocyte lysate (LAL) test. However, a nonspecific result is obtained, because the high concentration of serum proteins disturbs the enzyme reaction of the LAL test. In order to solve this problem, the LAL test was performed in this study after the centrifugation of the boiled serum samples to remove the impurities. As a result, among the various conditions examined, endotoxin measurement with the LAL test was the most accurate and repeatable after centrifugation of the boiled serum at $100^{\circ}C$. Moreover, the endotoxin was accurately and repeatedly measured from the prepared sera of mice that had been administered an intraperitoneal injection of purified lipopolysaccharides (LPS) or E. coli. Therefore, the application of centrifugation to remove impurities from boiled serum gives an accurate measurement of endotoxins in the sera of normal subjects or patients, and this will lead to the improved diagnosis and prevention of diseases caused by endotoxins. In addition, the centrifugation of boiled serum samples should be considered and included in the development of endotoxin test kits.

• Asian-Australasian Journal of Animal Sciences
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• v.19 no.8
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• pp.1095-1099
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• 2006

Leptin and IGFBP-3 are two proteins that play an important role in growth and metabolism of the animals. They are also involved in the immune function of animals and, thus, are candidate genes for the study of association with immune functions. Polymerase Chain Reaction-Restriction Fragment Length Polymorphism (PCR-RFLP) of these two genes was done to screen 64 crossbred (Holstein Friesian${\times}$Hariana) female calves of one year of age. From each RFLPs (fragments) three genotypes were observed. In all the RFLPs the mutant homozygotes were very less in numbers and, hence, were excluded from the least squares analysis. The serum IgG level was estimated using SRID assay. The mean level of serum IgG was $28.83{\pm}2.73mg/ml$. The effect of these identified genotypes on serum IgG level of calves at one year of age was analysed using least squares analysis. The HaeIII RFLP-AB genotype had significantly (p<0.05) higher serum IgG level ($31.86{\pm}3.05$) than the HaeIII RFLP-AA ($25.62{\pm}2.96$) genotype. There was no significant effect of leptin genotypes on the IgG level. The present results indicated a role of the IGFBP-3 gene on serum IgG level of cattle calves.

• Journal of Periodontal and Implant Science
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• v.45 no.1
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• pp.14-22
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• 2015

Purpose: The purpose of this study was to compare serum amyloid A (SAA) protein levels with high-sensitive C-reactive protein (hs-CRP) levels as markers of systemic inflammation in patients with chronic periodontitis. The association of serum titers of antibodies to periodontal microbiota and SAA/hs-CRP levels in periodontitis patients was also studied. Methods: A total of 110 individuals were included in this study. Patients were assessed for levels of hs-CRP and SAA. Nonfasting blood samples were collected from participants at the time of clinical examination. The diagnosis of adipose tissue disorders was made according to previously defined criteria. To determine SAA levels, a sandwich enzyme-linked immunosorbent assay was utilized. Paper points were transferred to a sterile tube to obtain a pool of samples for polymerase chain reaction processing and the identification of Porphyromonas gingivalis, Aggregatibacter actinomycetemcomitans, and Tannerella forsythia. The serum level of IgG1 and IgG2 antibodies to P. gingivalis, A. actinomycetemcomitans, and T. forsythia was also determined. Results: SAA and hs-CRP levels were higher in periodontitis patients than in controls (P<0.05). In bivariate analysis, high levels of hs-CRP (>3 mg/L) and SAA (>10 mg/L) were significantly associated with chronic periodontitis (P=0.004). The Spearman correlation analysis between acute-phase proteins showed that SAA positively correlated with hs-CRP (r=0.218, P=0.02). In the adjusted model, chronic periodontitis was associated with high levels of SAA (odds ratio [OR], 5.5; 95% confidence interval [CI], 1.6-18.2; P=0.005) and elevated hs-CRP levels (OR, 6.1, 95% CI, 1.6-23.6; P=0.008). Increased levels of serum IgG2 antibodies to P. gingivalis were associated with high levels of SAA (OR, 3.6; 95% CI, 1.4-8.5; P=0.005) and high concentrations of hs-CRP (OR, 4.3; 95% CI, 1.9-9.8; P<0.001). Conclusions: SAA and hs-CRP concentrations in patients with chronic periodontitis are comparably elevated. High serum titers of antibodies to P. gingivalis and the presence of periodontal disease are independently related to high SAA and hs-CRP levels.

• The Journal of Korean Society of Virology
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• v.27 no.2
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• pp.105-113
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• 1997

The truncated $E1_{192-283}$ and $E2_{384-649}$ genes of hepatitis C virus (HCV) linked to the gene for glutathione S-transferase (GST) were constructed and their expressions were analyzed. The $GST-E1_{192-283}$ fusion gene overexpressed the fusion protein in E. coli as a soluble form, while the $GST-E1_{192-383}$ plasmid did not express expected fusion protein. The purified $GST-E1_{192-283}$ fusion protein was efficiently cleaved by thrombin. More than 90% pure, HCV $E1_{192-283}$ protein was obtained by GST-agarose chromatography. The truncated $GST-E2_{384-649}$ fusion gene expressed the fusion protein mainly as an insoluble form, whereas the $GST-E2_{384-740}$ did not express the fusion protein. The truncated $GST-E1_{182-283}$ and $GST-E2_{384-649}$ fusion proteins reacted specifically with an HCV patient serum. In addition, mice immunized with either the purified $E1_{192-283}$ or $GST-E2_{384-649}$ proteins generated specific antibodies to each antigen. The results suggested that hydrophobic carboxyl portions of the E1 and E2 proteins might affect expression levels as well as the solubility of each fusion protein in bacteria. Also, the truncated E1 protein with Tyr-192 to Ser-283 contained antigenic epitope(s) which could be specifically recognized by an HCV patient serum.

• Journal of Veterinary Clinics
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• v.21 no.1
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• pp.20-22
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• 2004

Bacterial infection of canine atopic dermatitis is largely caused by Staphylococcus intermedius and may be a superficial or deep pyoderma. The Purpose of this study was to identify the major proteins of S. intermedius cell surface components in humoral immune response of atopic dermatitis dog. Sera samples were obtained from dogs with atopic dermatitis and superficial pyoderma referred to the Veterinary Medical Teaching Hospital, Konkuk University. An isolate of S. intermedius from a clinical case of canine atopic dermatitis was cultured in brain heart infusion broth overnight at $37^{\circ}C$ in aerobic conditions on an orbital shaker. Following culture, Staphylococci were harvested by centrifugation, washed in PBS, and resuspended in PBS containing lysostaphin. The soluble components were separated by centrifugation and were collected. The soluble extract of S. intermedius was separated by sodium dodecylsulphate polyacrylamide gel electrophoresis (SDS-PAGE). The proteins were electrophoretically transferred onto nitrocellulose membrane. Western blotting for the specificity of serum IgG antistaphylococcal antibody was performed with anti-dog-IgG and sera obtained from an atopic dermatitis case and a normal dog. The molecular masses of four major proteins of S. intermedius recognized by serum obtained from an atopic dermatitis case were 18, 31, 75, and 110 kDa as determined by Western blot analysis. The present study indicates that most dogs of S. intermedius infection with atopic dermatitis could have a significant humoral immune response to bacterial proteins of the causative organism.