• Smart Structures and Systems
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• 제8권1호
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• pp.17-37
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• 2011

The microcantilever (MCL) sensor is one of the most promising platforms for next-generation label-free biosensing applications. It outperforms conventional label-free detection methods in terms of portability and parallelization. In this paper, an overview of recent advances in our understanding of the coupling between biomolecular interactions and MCL responses is given. A dual compact optical MCL sensing platform was built to enable biosensing experiments both in gas-phase environments and in solutions. The thermal bimorph effect was found to be an effective nanomanipulator for the MCL platform calibration. The study of the alkanethiol self-assembly monolayer (SAM) chain length effect revealed that 1-octanethiol ($C_8H_{17}SH$) induced a larger deflection than that from 1-dodecanethiol ($C_{12}H_{25}SH$) in solutions. Using the clinically relevant biomarker C-reactive protein (CRP), we revealed that the analytical sensitivity of the MCL reached a diagnostic level of $1{\sim}500{\mu}g/ml$ within a 7% coefficient of variation. Using grazing incident x-ray diffractometer (GIXRD) analysis, we found that the gold surface was dominated by the (111) crystalline plane. Moreover, using X-ray photoelectron spectroscopy (XPS) analysis, we confirmed that the Au-S covalent bonds occurred in SAM adsorption whereas CRP molecular bindings occurred in protein analysis. First principles density functional theory (DFT) simulations were also used to examine biomolecular adsorption mechanisms. Multiscale modeling was then developed to connect the interactions at the molecular level with the MCL mechanical response. The alkanethiol SAM chain length effect in air was successfully predicted using the multiscale scheme.

• Journal of Microbiology and Biotechnology
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• 제21권5호
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• pp.529-535
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• 2011

Single-chain variable fragment (scFv) is a fusion protein of the variable regions of the heavy (VH) and light (VL) chains of immunoglobulin, connected with a short linker peptide of 10 to about 20 amino acids. In this study, the scFv of a monoclonal antibody against the third domain of human CD4 was cloned from OKT4 hybridoma cells using the phage display technique and produced in E. coli. The expression, production, and purification of anti-CD4 scFv were tested using SDS-PAGE and Western blot, and the specificity of anti-CD4 scFv was examined using ELISA. A 31 kDa recombinant anti-CD4 scFv was expressed and produced in bacteria, which was confirmed by SDS-PAGE and Western blot assays. Sequence analysis proved the ScFv structure of the construct. It was able to bind to CD4 in quality ELISA assay. The canonical structure of anti-CD4 scFv antibody was obtained using the SWISS_MODEL bioinformatics tool for comparing with the scFv general structure. To the best of our knowledge, this is the first report for generating scFv against human CD4 antigen. Engineered anti-CD4 scFv could be used in immunological studies, including fluorochrome conjugation, bispecific antibody production, bifunctional protein synthesis, and other genetic engineering manipulations. Since the binding site of our product is domain 3 (D3) of the CD4 molecule and different from the CD4 immunological main domain, including D1 and D2, further studies are needed to evaluate the anti-CD4 scFv potential for diagnostic and therapeutic applications.

• Korean Journal of Veterinary Research
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• 제43권1호
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• pp.129-137
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• 2003

Porcine reproductive and respiratory syndrome (PRRS) is characterized by reproductive failures in sows and respiratory problems in piglets. The nucleocapsid(N) protein, encoded by the open reading frame 7 (ORF7) gene, is known to be the most abundant and antigenic protein in PRRS virus. Therefore, it was suggested that the N protein could be a suitable candidate for the detection of PRRS virus-specific antibodies and diagnosis of PRRS. In the present study, the ORF7 gene encoding the N protein was cloned and expressed as a fusion protein with the glutathione S-transferase (GST) in Escherichia coli. The resulting GST-N recombinant protein was used as an antigen for an indirect sandwich enzyme-linked immunosorbent assay (i-ELISA). Expressed GST-N recombinant protein was migrated at 41 kDa and reacted with ORF7-specific monoclonal antibody by Western blotting. In order to increase the specificity of the ELISA for the detection of PRRS virus-specific antibodes, an i-ELISA was developed using an anti-GST antibody as a capture antibody. The sensitivity and specificity of developed i-ELISA were 92% and 96%, respectively. Based on these results, it was suggested that the i-ELISA is a simple and rapid test for screening a large number of swine sera for the anti-PRRS virus antibodies.

• Asian Pacific Journal of Cancer Prevention
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• 제14권3호
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• pp.1635-1642
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• 2013

Helicobacter pylori antigen was prepared from an isolate from a patient with a duodenal ulcer. Serum samples were obtained from culture-positive H. pylori infected patients with duodenal ulcers, gastric ulcers and gastritis (n=30). As controls, three kinds of sera without detectable H. pylori IgG antibodies were used: 30 from healthy individuals without history of gastric disorders, 30 from patients who were seen in the endoscopy clinic but were H. pylori culture negative and 30 from people with other diseases. OFF-GEL electrophoresis, SDS-PAGE and Western blots of individual serum samples were used to identify protein bands with good sensitivity and specificity when probed with the above sera and HRP-conjugated anti-human IgG. Four H. pylori protein bands showed good (${\geq}$ 70%) sensitivity and high specificity (98-100%) towards anti-Helicobacter IgG antibody in culture-positive patients sera and control sera, respectively. The identities of the antigenic proteins were elucidated by mass spectrometry. The relative molecular weights and the identities of the proteins, based on MALDI TOF/TOF, were as follows: CagI (25 kDa), urease G accessory protein (25 kDa), UreB (63 kDa) and proline/pyrroline-5-carboxylate dehydrogenase (118 KDa). These identified proteins, singly and/or in combinations, may be useful for diagnosis of H. pylori infection in patients.

• IMMUNE NETWORK
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• 제10권2호
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• pp.35-45
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• 2010

Background: Expression of recombinant antibodies and their derivatives fused with other functional molecules such as alkaline phosphatase in Escherichia coli is important in the development of molecular diagnostic reagents for biomedical research. Methods: We investigated the possibility of applying a well-known Fos-Jun zipper to dimerize $V_H$ and $V_L$ fragments originated from the Fab clone (SP 112) that recognizes pyruvate dehydrogenase complex-E2 (PDC-E2), and demonstrated that the functional zipFv-112 and its alkaline phosphatase fusion molecules (zipFv-AP) can be produced in the cytoplasm of Origami(DE3) trxB gor mutant E. coli strain. Results: The zipFv-AP fusion molecules exhibited higher antigen-binding signals than the zipFv up to a 10-fold under the same experimental conditions. However, conformation of the zipFv-AP seemed to be influenced by the location of an AP domain at the C-terminus of $V_H$ or $V_L$ domain [zipFv-112(H-AP) or zipFv-112(L-AP)], and inclusion of an AraC DNA binding domain at the C-terminus of VH of the zipFv-112(L-AP), termed zipFv-112(H-AD/L-AP), was also beneficial. Cytoplasmic co-expression of disulfide-binding isomerase C (DsbC) helped proper folding of the zipFv-112(H-AD/L-AP) but not significantly. Conclusion: We believe that our zipFv constructs may serve as an excellent antibody format bi-functional antibody fragments that can be produced stably in the cytoplasm of E. coli.

• Molecular & Cellular Toxicology
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• 제2권4호
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• pp.273-278
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• 2006

95 squamous cell lung carcinoma samples (normal tissue: 40 samples, tumor: 55 samples) were analyzed with 8 K cDNA microarray. 1-way ANOVA test was employed to select differentially expressed genes in tumor with FDR<0.01. Among the selected 1,655 genes, final 212 genes were chosen according to the expression fold change and used for following analysis. The expression of up-regulated 64 genes was verified with Reverse Transcription PCR and 10 genes were identified as candidates for SCC markers. In our opinion, those candidates can be exploited as diagnostic or therapeutic purposes. Gene Ontology (GO) based analysis was performed using those 212 genes, and following categories were revealed as significant biological processes: Immune response (GO: 0006955), antigen processing (GO: 0030333), inflammatory response (GO: 0006954), Cell adhesion (GO: 0007155), and Epidermis differentiation (GO: 0008544). Gene set enrichment analysis (GSEA) also carried out on overall gene expression profile with 522 functional gene sets. Glycolysis, cell cycle, K-ras and amino acid biosynthesis related gene sets were most distinguished. These results are consistent with the known characteristics of SCC and may be interconnected to rapid cell proliferation. However, the unexpected results from ERK activation in squamous cell carcinoma gripped our attention, and further studies are under progress.

• Journal of Yeungnam Medical Science
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• 제16권2호
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• pp.369-375
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• 1999

Chordoma is a slow-growing malignant neoplasm arising from the remnants of the primitive notochord. It accounts for 1 to 4% of all malignant bone tumors. It occurs exclusively along the spinal axis. Authors experienced four cases of chordoma occurred in the sacrococcygeal region. There were two male and two female patients, with a mean age of 63.5 years(range, 57~75 years). Tissue was obtained by wide excision in two patients, by incisional biopsy in one patient and by needle biopsy in the other. Adjuvant radiation therapy was performed on all the patients after their biopsy. The mean diameter of the tumors was 7.6cm(range, 5.5 to 13.0cm). Grossly, tumor was multiobulated, soft and myxoid gelatinous mass. Microscopically, the tumor showed lobulated feature divided by fibrous septa within it. There were physaliphorous cells with vacuolated bubbly cytoplasm. And small uniform, round, and non-vacuolated tumor cells were also present. On immunohistochemical stain, all the cases were immunoreactive for cytokeratin, epithelial membrane antigen(EMA) and vimentin, respectively. One of the 4 cases was positive for S-100 protein. All the cases were negative for CEA.

• Journal of Preventive Medicine and Public Health
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• 제10권1호
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• pp.150-154
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• 1977

In the diagnosis of parasitic helminthic diseases, the value of examining and identifying the parasitic eggs and/or adult parasites from patient's urine or stool are well appreciated. However, these methods have a limited value in the diagnosis of tissue or intracellular parasitism, and we have to rely on supplementary methods such as immune-serological test. The author tested the value of gel precipitation reactions as a diagnostic method of clonorchiasis by observing the appearance of bands in rats experimentally infected with Clonorchis sinensis, And the therapeutic effect of CIBA 35'058-Ba was evaluated by this serological method. The antigen was prepared from the adult worms infected in rabbits by Tsuji method. Rats infected with 40 metacercariae each were bled on 7,14,21,26,28,39,42(43),49(53) days after infection to observe the appearance of precipitin bands by both Oucterlony method and immunoelectrophoresis. Fifteen rats were separately infected and treated with CIBA 35'058-Ba in dose of 15mg/kg of body weight. The following results were obtained: 1. It was observed that there exist individual variations in the appearance of the first precipitin band with the range of 2-4 weeks after infection. 2. The number of precipitin bands was increased until 6-7 weeks after infection. In all cases, 3 precipitin bands were appeared by Oucterlony method and 6-7 bands were appeared by immunoelectrophoresis after 6-7 weeks of infecion. 3. It was hardly possible to notice any change in the number of bands after the administration of CIBA 35'058-Ba. This result suggested that the drug has no effect on clonorchiasis which was confirmed by the autopsy of the experimental rats.

• Tuberculosis and Respiratory Diseases
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• 제41권1호
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• pp.36-41
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• 1994

Angiosarcoma is a very rare malignant tumor of endothelial cell origin. We experienced a case of angiosarcoma presented with massive pleural effusion, which was considered as a metastasis from right kidney. A 44-year-old male patient was admitted due to dyspnea for one month. He had a history of transient hematuria 3 months before admission, which disappeared spontaneously. Chest roentgenography showed total haziness in left hemithorax with multiple nodular shadows in right lung. Abdominal ultrasonogram showed a single heterogeneous hyperechoic mass, measuring about $7.3{\times}7.1{\times}6.5cm$ in size in the upper and mid-pole of the right kidney, involving renal sinus. Computed tomography of the chest revealed highly enhanced multiple pulmonary and subpleural nodules with loculated pleurisy. In bronchoscopic finding, a fungating, hypervascular tumor mass was noticed at the orifice of anterior basal segment of left lower lung after removal of tenaceous mucus. Pleural and bronchoscopic biopsies showed findings of angiosarcoma confirmed by immunochemical stains with factor VIII related antigen(+), laminin(+) and vimentin(+), and by characteristic electronmicroscopic findings. Massive pleural effusion was controlled with several times of pleurodesis in both pleural spaces.

• Asian Pacific Journal of Cancer Prevention
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• 제16권12호
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• pp.4891-4894
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• 2015

Background: To evaluate the value of combined detection of serum carcinoembryonic antigen (CEA), cytokeratin 19 fragment (CYFRA21-1), and carbohydrateantigen 125 (CA125) for the clinical diagnosis of nonsmall cell lung cancer (NSCLC). Materials and Methods: Serum CEA, CYFRA21-1 and CA125 were assessed in 140 patients with NSCLC, 90 patients with benign lung disease and 90 normal control subjects, and differences of expression were compared in each group, and joint effects of these tumor markers in the diagnosis of NSCLC were analyzed. Results: Serum CEA, CYFRA21-1 and CA125 in patients with NSCLC were significantly higher than those with benign lung disease and normal controls (P<0.05). The sensitivity of CEA, CYFRA21-1 and CA125 were 49.45%, 59.67%, and 44.87% respectively. As expected, combinations of these tumor markers improved their sensitivity for NSCLC. The combined detection of CEA + CYFRA21-1 was the most cost-effective combination which had higher sensitivity and specificity in NSCLC. Elevation of serum CEA and CYFRA21-1 was significantly associated with pathological types (P<0.05) and elevation of serum CEA, CYFRA21-1 and CA125 was significantly associated with TNM staging (P<0.05). Conclusions: Single measurement of CEA, CYFRA21-1 and CA125 is of diagnostic value in the diagnosis of lung cancer, and a joint detection of these three tumor markers, could greatly improve the sensitivity of diagnosis on NSCLC. Combined detection of CEA + CYFRA21-1 proved to be the most economic and practical strategy in diagnosis of NSCLC, which can be used to screen the high-risk group.