• Korean Journal of Microbiology
• /
• v.52 no.1
• /
• pp.10-17
• /
• 2016

Enzyme immunoassay to analyze specific binding activity of antibody to antigen uses horseradish peroxidase (HRP) or alkaline phosphatase (AP). Chemical methods are usually used for coupling of these enzymes to antibody, which is complicated and random cross-linking process. As results, it causes decreases or loss of functional activity of either antibody or enzyme. In addition, most enzyme assays use secondary antibody to detect antigen binding activity of primary antibody. Enzymes coupled to secondary antibody provide a binding signal by substrate-based color development, suggesting secondary antibody is required in enzyme immunoassay. Additional incubation time for binding of secondary antibody should also be necessary. More importantly, non-specific binding activity caused by secondary antibody should also be eliminated. In this study, we cloned AP isolated from Escherichia coli (E. coli) chromosome by PCR and fused to) hAY4 single-chain variable domain fragment (ScFv) specific to death receptor (DR4) which is a receptor for tumor necrosis factor ${\alpha}$ related apoptosis induced ligand (TRAIL). hAY4 ScFv-AP expressed in E. coli showed 73.8 kDa as a monomer in SDS-PAGE. However, this fusion protein shown in size-exclusion chromatography (SEC) exhibited 147.6 kDa as a dimer confirming that natural dimerization of AP by non-covalent association induced ScFv-AP dimerization. In several immunoassay such as ELISA, Western blot and immunocytochemistry, it showed antigen binding activity by color development of substrates catalyzed by AP directly fused to primary hAY4 ScFv without secondary antibody. In summary, hAY4 ScFv-AP fusion protein was successfully purified as a soluble dimeric form in E. coli and showed antigen binding activity in several immunoassays without addition of secondary antibody which sometimes causes time-consuming, expensive and non-specific false binding.

• Korean Journal of Food Science and Technology
• /
• v.31 no.1
• /
• pp.238-245
• /
• 1999

Antioxidative effects of the water or ethanol extracts from Rhus verniciflua Stokes (RVS) were measured by protection against hydroxyl radicals in mouse brain tissue culture. In the water extracts from RVS, cell viabilities were estimated 60.0, 66.0, 72.0, 84.0 and 90.0% at addition of 1, 2, 4, 7 and $10{\mu}L$, respectively, compared with GO (20 mU/mL) alone. The cell viability in the ethanol extracts was similarly with water extracts. In the antitumor effects, the results showed that percentages of the HeLa cell death were approximately 24% for 12 hrs, 57% for 48 hrs at addition of 10%/well ethanol extracts respectively. To know inhibition of tumor growth, in vivo, mice (BALB/c) were inoculated with 0.25 mL CT-26 $(1{\times}10^6\;cells/mL)$ subcutaneously. After the generation of tumor, the results of RVS extracts (ethanol, water) injection showed generally that the tumor size in BALB/c was reduced. For physicochemical characterization of the RVS extracts, purified substances of water or ethanol extracts were analized with SDS-PAGE and ICP spectrometer. In electrophoresis, gel showed 2 bands (210, 230 KDa). The results of ICP verified that RVS extracts contain $Cu^{2+}$ in both samples. Conclusively, this substance might be a laccase which has a biological effective function, as a natural bioactive substance.

• Pediatric Infection and Vaccine
• /
• v.18 no.2
• /
• pp.182-192
• /
• 2011

Purpose : Atopic findings may be associated with severity of pneumonia in 2009 pandemic influenza A (H1N1) infection, which could suggest a possible association between atopic findings and the severity of viral infections. Thus, we studied association between atopic findings and severity of disease in children with H1N1 influenza infection. Methods : A retrospective study was performed in 74 children admitted in a single tertiary institute and confirmed as H1N1 patients by reverse transcriptase (RT) - polymerase chain reaction (PCR). They were divided into 2 groups according to the severity of pneumonia. We evaluated whether the atopic finding is risk factor between the two groups. Results : Children with severe pneumonia had higher percentages of serum eosinophilia (88% vs 40%, P <0.001), asthma (65% vs 35%, P =0.011), allergic rhinitis (71% vs 40%, P =0.009), and IgE level (P =0.007). We found positive correlations between aeroallergen sensitizations and severity of pneumonia (82% vs 53%, P =0.007). Conclusion : Among patients with H1N1 pneumonia, asthma and atopic findings are risk factors for severity of pneumonia.

• Nuclear Medicine and Molecular Imaging
• /
• v.42 no.4
• /
• pp.267-274
• /
• 2008

Purpose: Transient neurological deterioration (TND) is one of the complications after extracranial-intracranial bypass surgery, and it has been assumed to be caused by postoperative transient hyperperfusion. This study was performed to evaluate the relationship between TND and preoperative and postoperative cerebral perfusion status on brain perfusion SPECT following superficial temporal artery - middle cerebral artery (STA-MCA) anastomosis surgery. Materials and Methods: A total of 60 STA-MCA anastomosis surgeries of 56 patients (mean age: $50{\pm}16$ yrs; M:F=29:27; atherosclerotic disease: 33, moyamoya disease: 27) which were done between September 2003 and July 2006 were enrolled. The resting cerebral perfusion and cerebral vascular reserve (CVR) after acetazolamide challenge were measured before and 10 days after surgery using 99mTc-ethylcysteinate dimer (ECD) SPECT. Moreover, the cerebral perfusion was measured on the third postoperative day. With the use of the statistical parametric mapping and probabilistic brain atlas, the counts for the middle cerebral artery (MCA) territory were calculated for each image, and statistical analyses were performed. Results: In 6 of 60 cases (10%), TND occurred after surgery. In all patients, the preoperative cerebral perfusion of affected MCA territory was significantly lower than that of contralateral side (p=0.002). The cerebral perfusion on the third and tenth day after surgery was significantly higher than preoperative cerebral perfusion (p=0.001, p=0.02). In TND patients, basal cerebral perfusion and CVR on preoperative SPECT were significantly lower than those of non-TND patients (p=0.01, p=0.05). Further, the increases in cerebral perfusion on the third day after surgery were significant higher than those in other patients (p=0.008). In patients with TND, the cerebral perfusion ratio of affected side to contralateral side on third postoperative day was significantly higher than that of other patients (p=0.002). However, there was no significant difference of the cerebral perfusion ratio on preoperative and tenth postoperative day between patients with TND and other patients. Conclusion: In patients with TND, relative and moderate hyperperfusion was observed in affected side after bypass surgery. These finding may help to understand the pathophysiology of TND.

• Tuberculosis and Respiratory Diseases
• /
• v.44 no.1
• /
• pp.44-51
• /
• 1997

Background : The changes of the composition in the T-lymphocyte are important as an immunological abnormality in the pathogenesis of tuberculosis. Previously, the second type of TCR dimer(${\gamma}{\delta}$ T lymphocyte) that did not express CD4 or CD8 molecules was found. In other reports the presence of this type of lymphocytes was increased in the initial stage of tuberculous infections. Method : To determine whether there are some differences in the T-lymphocyte subsets in the peripheral blood or pleural effusion between pleural tuberculosis and other pleurisy. Thirty patients with pleural effusion among the forty-nine patients were examined T-lymphocyte subset analysis(CD4+T-cell,CD8+ T-cell,${\gamma}{\delta}$ T-lymphocytes) with anti- Leu4, anti-Leu3a, anti-Lea2a, anti HLA-DR and anti-TCR-${\gamma}{\delta}$-1(Becton & Dickinson Co.). Results : The average age of the patients was 50 years old(17-81year). There were 33 males and 16 female patients. Patiensts with tuberculosis are 30cases(tuberculous pleurisy 15), lung cancer 12cases(malignant effusion 9) and pneumonia 7cases(parapneumonic effusion 6cases) In T lymphocyte subsets of pleural effusion, helper T lymphocyte(54.6 + 13.8 %) of tuberculous pleurisy was higher than that(36.2 + 25.3 %) of non-tuberculous pleurisy(p=0.04). The peripheral blood ${\gamma}{\delta}$ T-lymphocytes in tuberculousis was insignificantly higher than non-tuberculous patients(p= 0.24). The peripheral blood ${\gamma}{\delta}$ T-lymphocytes and pleural ${\gamma}{\delta}$ T-Iymphocytes in tuberculous pleurisy was insignificantly higher than in non-tuberculous pleurisy(p= 0.16, p= 0.12). Conclusion : The percentage of -${\gamma}{\delta}$ T lymphocytes among the total T-lymphocytes is not significantly increased in the peripheral blood or pleural effusion of the pleural tuberculosis. ${\gamma}{\delta}$ T lymphocytes is less useful as a diagnostic method of pleural tuberculosis.

• The Korean Journal of Nuclear Medicine
• /
• v.32 no.6
• /
• pp.490-496
• /
• 1998

Purpose: We evaluated brain perfusion SPECT findings of MELAS syndrome and mitochondrial myopathy in correlation with MR imaging in search of specific imaging features. Materials and Methods: Subjects were five patients (four females and one male; age range, 1 to 25 year) who presented with repeated stroke-like episodes, seizures or developmental delay or asymptomatic but had elevated lactic acid in CSF and serum. Conventional non-contrast MR imaging and Tc-99m-ethyl cysteinate dimer (ECD) brain perfusion SPECT were Performed and imaging features were analyzed. Results: MRI demonstrated increased T2 signal intensities in the affected areas of gray and white matters mainly in the parietal (4/5) and occipital lobes (4/5) and in the basal ganglia (1/5), which were not restricted to a specific vascular territory. SPECT demonstrated decreased perfusion in the corresponding regions of MRI lesions. In addition, there were perfusion defects in parietal (1 patient), temporal (2), and frontal (1) lobes and basal ganglia (1) and thalami (2). In a patient with mitochondrial myopathy who had normal MRI, decreased perfusion was noted in left parietal area and bilateral thalami. Conclusion: Tc-99m ECD SPECT imaging in patients with MELAS syndrome and mitochondrial myopathy showed hypoperfusion of parieto-occipital cortex, basal ganglia, thalamus and temporal cortex, which were not restricted to a specific vascular territory. There were no specific imaging features on SPECT. The significance of abnormal perfusion on SPECT without corresponding MR abnormalities needs to be evaluated further in larger number of patients.

• The Korean Journal of Nuclear Medicine
• /
• v.36 no.4
• /
• pp.244-254
• /
• 2002

Purpose: This study investigated the differences between technetium-99m ethyl cysteinate dimer (Tc-99m ECD) and technetium-99m hexamethylpropylene amine oxime (Tc-99m HMPAO) uptake in the normal brain by means of statistical parametric mapping (SPM) analysis. Materials and Methods: We retrospectively analyzed age and sex matched 53 cases of normal brain SPECT. Thirty-two cases were obtained with Tc-99m ECD and 21 cases with Tc-99m HMPAO. There were no abnormal findings on brain MRIs. All of the SPECT images were spatially transformed to standard space, smoothed and globally normalized. The differences between the Tc-99m ECD and Tc-99m HMPAO SPECT images were statistically analyzed using statistical parametric mapping (SPM'99) software. The differences bgetween the two groups were considered significant ant a threshold of corrected P values less than 0.05. Results: SPM analysis revealed significantly different uptakes of Tc-99m ECD and Tc-99m HMPAO in the normal brains. On the Tc-99m ECD SPECT images, relatively higher uptake was observed in the frontal, parietal and occipital lobes, in the basal ganglia and thalamus, and in the superior region of the cerebellum. On the Tc-99m HMPAO SPECT images, relatively higher uptakes was observed in subcortical areas of the frontal region, temporal lobe, and posterior portion of inferior cerebellum. Conclusion: Uptake of Tc-99m ECD and Tc-99m HMPO in the normallooking brain was significantly different on SPM analysis. The selective use of Tc-99m ECD of Tc-99m HMPAO in brain SPECT imaging appears especially valuable for the interpretation of cerebral perfusion. Further investigation is necessary to determine which tracer is more accurate for diagnosing different clinical conditions.