• Biomedical Science Letters
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• v.9 no.2
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• pp.67-74
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• 2003

Viral and non-viral vectors have been used in the delivery of genetic materials into animal cells and tissues, with each approach having pros and cons. Non-viral vectors have many useful merits such as easy preparation, low immunity and size tolerance of a transgene when compared to those of viral vectors. Delivery specificity may be achieved by complex formation between receptor ligands and a non-viral vector. In the present study, non-viral vector systems are investigated in an effort to find a practical delivery means for gene therapy, Receptor-ligand interaction between transferrin-receptor and transferrin was utilized for efficient gene transfer into cancer cells. A plasmid vector, pcDNA3 (LacZ) was ligated with a small duplexed oligo fragment in which a Biotin- VN$^{TM}$ phosphoramidite was placed in the middle of the oligo. The plasmid vector labeled by biotin was then conjugated with biotin-labeled transferrin via streptavidin. This trimeric conjugates were delivered to a hepatoma cell line, HepG2. The delivery efficiency of the trimeric conjugate was 2-fold higher than that of cationic liposomes used for transfection of a plasmid vector. These results demonstrate that a plasmid vector can be efficiently transferred into cells by forming a trimeric complex of plasmid vector-linker-ligand.

• Journal of Periodontal and Implant Science
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• v.46 no.1
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• pp.2-9
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• 2016

Purpose: Porphyromonas gingivalis and Tannerella forsythia have been implicated as the major etiologic agents of periodontal disease. These two bacteria are frequently isolated together from the periodontal lesion, and it has been suggested that their interaction may increase each one's virulence potential. The purpose of this study was to identify proteins on the surface of these organisms that are involved in interbacterial binding. Methods: Biotin labeling of surface proteins of P. gingivalis and T. forsythia and liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis was performed to identify surface proteins involved in the coaggregating activity between P. gingivalis and T. forsythia. Results: It was found that three major T. forsythia proteins sized 161, 100, and 62 kDa were involved in binding to P. gingivalis, and P. gingivalis proteins sized 35, 32, and 26 kDa were involved in binding to T. forsythia cells. Conclusions: LC-MS/MS analysis identified one T. forsythia surface protein (TonB-linked outer membrane protein) involved in interbacterial binding to P. gingivalis. However, the nature of other T. forsythia and P. gingivalis surface proteins identified by biotin labeling could not be determined. Further analysis of these proteins will help elucidate the molecular mechanisms that mediate coaggregation between P. gingivalis and T. forsythia.

• Journal of Physiology & Pathology in Korean Medicine
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• v.18 no.4
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• pp.1154-1158
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• 2004

The classical ABC (avidin-biotin peroxidase complex) method for immunohistochemistry in the paraffin-embedded tissues bring into being disadvantage such as low sensitivity of antigen detection and highly background. The biotinyl-tyramide conjugation recently introduced for sensitive immunohistochemistry was applied to light microscopy in paraffin-embedded pancreatic and liver tissues. The protocol consists of an indirect method in which 4-5㎛ tissue sections are reacted successively within a specific primary antibody, followed by a biotinylated secondary antibody, streptavidin-horseradich peroxidase (HRP), and then finally with biotinyl-tyramide. The labeling obtained for insulin and collagen antigen tested in pancreatic and liver tissues, respectively, was found to be highly specific with the labeling for each antigen confined to its particular cellular compartment. In this study, fish (flounder) serum was specially applied to remove nonspecific binding. Background levels and nospecific deposition of the staining were negligible. This results suggest that super-signal induction method by biotinyl-tyramide conjugate can readily applied to antigen detection of the paraffin-embedded tissues.

• Journal of Food Hygiene and Safety
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• v.31 no.5
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• pp.327-334
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• 2016

This study was conducted to establish the standard method for the contents of biotin in milk formulas. To optimize the method, we compared several conditions for liquid extraction, purification and instrumental measurement using spiked samples and certified reference material (NIST SRM 1849a) as test materials. LC-MS/MS method for biotin was established using $C_{18}$ column and binary gradient 0.1% formic acid/acetonitrile, 0.1% formic acid/water mobile phase is applied for biotin. Product-ion traces at m/z 245.1 ${\rightarrow}$ 227.1, 166.1 are used for quantitative analysis of biotin. The linearity was over $R^2=0.999$ in range of $5{\sim}60{\mu}g/L$. For purification, chloroform was used as a solvent for eliminating lipids in milk formula. The linearity was over 0.999 in range of 5~60 ng/mL. The detection limit and quantification limit were 0.10, 0.31 ng/mL. The accuracy and precision of LC-MS/MS method using CRM were 103%, 2.5% respectively. Optimized methods were applied in sample analysis to verify the reliability. All the tested milk formulas were acceptable contents of biotin compared with component specification and standards for nutrition labeling. The standard operating procedures were prepared for biotin to provide experimental information and to strengthen the management of nutrient in milk formula.

• Proceedings of the Korean Institute of Electrical and Electronic Material Engineers Conference
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• 2010.06a
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• pp.280-280
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• 2010

Alzheimer disease is a progressive neurodegenerative disease of the aged, characterized by memory loss and dementia. For diagnosis of Alzheimer disease we have simply modified macrophage with amyloid beta bonded with different molecules. Modified Macrophage was observed with microscope for co-localization of amyloid beta molecule. For this experiment we used fluoroscene labeling substances. The macrophage was modified also with cell staining method. For cell staining method was used avidin-biotin reaction principles. All experiments were carried out on poly-L-lysine coated and sterilized glass substrates. In the presentation we will show the further investigations and applications with modified macrophage.

• Journal of Dairy Science and Biotechnology
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• v.33 no.4
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• pp.271-275
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• 2015

Colloidal semiconductor CdSe-ZnS core-shell nanocrystal quantum dot (Qdot) are luminescent inorganic fluorophores that show potential to overcome some of the functional limitations encountered with organic dyes in fluorescence labeling applications. Salmonella Enteritidis has emerged as a major cause of human salmonellosis worldwide since the 1980s. A rapid, specific, and sensitive method for the detection of Salmonella Enteritidis was developed using Qdot as a fluorescence marker coupled with immunomagnetic separation. Magnetic beads coated with anti-Salmonella Enteritidis antibodies were employed to selectively capture the target bacteria, and biotin-conjugated anti-Salmonella antibodies were added to form sandwich immune complexes. After magnetic separation, the immune complexes were labeled with Qdot via biotin-streptavidin conjugation, and fluorescence measurement was carried out using a fluorescence measurement system. The detection limit of the Qdot method was a Salmonella Enteritidis concentration of $10^3$ colony-forming units (CFU)/mL, whereas the conventional fluorescein isothiocyanate-based method required over $10^5CFU/mL$. The total detection time was within 2 h. In addition to the potential for general nanotechnology development, these results suggest a new rapid detection method of various pathogenic bacteria from a complex food matrix.

• Proceedings of the Korean Vacuum Society Conference
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• 2011.08a
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• pp.389-389
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• 2011

최근 생체분자 구조 연구가 의료진단, 생명 현상 규명 및 의약품 개발 등 다양한 분야에 응용되고 있으나 대부분의 분석방법이 제한적이어서 새로운 기술 개발의 필요성이 증대하고 있다. 종래의 DNA 등의 생체분자의 분석은 형광염료를 이용한 방법이 주로 이용되었다. 형광염료는 단백질을 포함한 여러 물질들에 대해 반응하지 않기 때문에 분석에 제한이 있으며, 이와 같은 단점을 보완하는 방법으로 SPR (surface plasmon resonance) 분석법이 연구되었다. SPR은 형광염료 분석에 필수적인 레이블링(labeling) 등의 전처리 과정 없이 높은 민감도로 분석이 가능한 장점이 있다. 한편, 그래핀은 뛰어난 전자기적 성질과 기계적 성질 을 가지는 반금속(semimetal)으로, 실험실 규모에서 안정적인 합성이 실현되면서 그 응용 분야에 대한 연구가 활발히 이루어 지고 있다. 그래핀은 큰 표면적 대 부피비를 가지며, 이는 검출물질과의 반응성이 좋아야 하는 센서기술에 있어서 장점으로 작용한다. 특히, 비금속성을 띠는 단층 그래핀을 여러 장 겹치면 금속성을 갖게 되기 때문에 SPR 센서의 금속 필름으로 응용이 가능하다. 본 연구에서는 SPR 현상을 이용하는 광섬유 센서의 감도와 정확도를 증진시키기 위해 광섬유 표면에 그래핀을 적용하였다. 광섬유는 상부 피복과 클래딩을 제거하여 코어를 노출시킨 후, 다층 그래핀 필름을 코팅함으로써 검출부를 구성한다. 그 후, DNA-biotin 용액, DNA-biotin 용액, 그리고 Streptavidin 단백질 복합 용액에 대한 검출기 신호를 분석하였다. 구성된 센서에 각 용액을 1 ${\mu}{\ell}$ 씩 반응시켜 분광계로 파장에 따른 광강도를 측정하는 실험을 수행했으며, 450 nm에서 460 nm 범위의 푸른빛의 광원을 사용하였다. 그래핀 필름의 유무에 따라 확연히 구분되는 경향을 보이는 결과를 얻었고 그래핀 필름이 기존 SPR 센서의 금속박막을 대체 할 수 있음을 확인하였다.

• Journal of Korean Neurosurgical Society
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• v.39 no.6
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• pp.432-437
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• 2006

Objective : In order to establish the index of degeneration, the authors performed a histochemical study with Safranin-O staining and investigated the occurrence of apoptosis in the human intervertebral disc. Methods : Eighteen intervertebral disc specimens surgically extracted from the patients and two additional specimens from the autopsied cases were stained with Safranin-O for proteoglycan according to a standard protocol. Terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate- biotin nick end labeling[TUNEL] was used to detect the fragmented DNA known to be associated with apoptotic cell death and classification scheme was formulated for categorization of the degree of Safranin-O staining [normal, moderate reduction, faint] by modification of Makin's histological-histochemical grading. The Kruskal-Wallis H test and Chi-square test were used for statistical analysis. Results : The statistical results showed a significant difference in the mean age between "normal" Safranin-O staining group and the others [19.3 versus 55, 43.4, p=0021]. However, there was no statistically significant correlation between Safranin-O staining and MR grading of disc degeneration. Only six of eighteen surgical specimens and none in autopsies showed positive apoptotic cells in TUNEL staining. Conclusion : The determination of the degree of degeneration in surgically obtained disc tissue per se by histochemical staining or by the degree of apoptosis that corresponds to its morphologic change was not feasible.

• Journal of Korean Neurosurgical Society
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• v.30 no.4
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• pp.430-436
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• 2001

Objective : The proliferative potential of intracranial glioma affects the histological malignancy and prognosis of patients with these tumors. In this study, we present the relationship between MIB-1 labeling index(LI) and clinical variables which might play the major role in determining the prognosis of patient with astrocytic tumors. Patients and Methods : Excised tumor specimens from a total of 52 patients were stained to detect monoclonal MIB-1-Ki-67 antibody by avidin-biotin complex immunohistochemistry. The MIB-1 LI was evaluated with histological grades, demograpghic data, and survival time. The statistical significance of their correlation was analyzed by Pearson correlation test. Results : The 52 patients included 30 male patients and 22 female patients. The tumors according to the criteria of the World Health Organization(WHO) classification were verified as pleomorphic xanthoastrocytoma in one, pilocytic astrocytomas 4, astrocytomas 1, anaplastic astrocytomas 3, and glioblastomas 31. MIB-1 LI in astrocytic ttumors showed no correlation with age and gender. However, the patients under 10 years had the longest survival time, whereas short survival time was observed in the older patients. The mean MIB-1 LI of different tumor grades were as follows : pleomorphic xanthoastrocytoma, $4.40{\pm}0.00$ ; pilocytic astrocytoma, $4.53{\pm}3.09$ ; astrocytoma, $5.50{\pm}6.03$ ; anaplastic astrocytoma, $12.68{\pm}12.50$ ; Glioblastoma, $21.31{\pm}19.63$. Although the levels of MIB-1 LI were varied in individual tumors, the MIB-1 LI was increased in parallel with the histological grades. Glioblstomas showed significantly higher MIB-1 LI compared with that of anaplastic astrocytomas and low grade astrocytomas (p = 0.001). The mean survival time of entire group of patients was also well correlated with MIB-1 LI in astrocytic tumors(p = 0.015). Moreover, the mean survival time of the entire group of patients with Lis < 10 was $125.33{\pm}113.57weeks$, and the mean survival of those with $Lis{\geq}10$ was $60.71{\pm}62.58weeks$. This difference was also statistically significant(p = 0.004). Conclusion : The results of this study suggest that MIB-1 LI correlates with histological grades and might play a significant role in predicting the survival of patients with astrocytic tumors.

• The Journal of Korean Medicine
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• v.27 no.3 s.67
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• pp.51-62
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• 2006

Objective: Although chronic non-bacterial prostatitis is a common disease, it is very difficult to treat effectively. Lygodium japonicum has traditionally been used in treatment of urinary tract inflammation and voiding disturbance. In this study, we investigated the therapeutic effects and action mechanism of Lygodium japonicum in the rat model of non-bacterial prostatitis induced by castration and testosterone treatment. Methods: Five-month-old rats were treated with $17\beta-estradiol$ after castration for induction of experimental non-bacterial prostatitis, which is similar to human chronic prostatitis in histopathological profiles. Lygodium japonicum and testosterone were administered as an experimental specimen and a positive control, respectively. The prostates were evaluated by histopathological parameters including the epithelial score and epithelio-stromal ratio for glandular damage, proliferating cell nuclear antigen (PCNA) labeling index for cyto-proliferation and a TUNEL (deoxyuridine triphosphate biotin nick end-labeling) assay for cell apoptosis. Results: While prostates of control rats revealed severe acinar gland atrophy and stromal proliferation, the rats treated with Lygodium japonicum showed a lesser range of tissue damage. Epithelial score was improved in Lygodium japonicum than that of the control (P<0.05). The epithelio-stromal ratio was lower in Lygodium japonicum when compared to that of the control (P<0.05). Although there was no difference in PCNA and TUNEL positive cells of the glandular epithelia, we found an decreased number of PCNA positive cell and concurrent increase of TUNEL positive cells in the stroma of Lygodium japonicum treated rats (P<0.01). Conclusions: These findings suggest that Lygodium japonicum may protect the glandular epithelial cells and also inhibit stromal proliferation in association with suppression of cyto-proliferation and stimulation of apoptosis. We concluded that Lygodium japonicum may be a useful remedy agent for treating the chronic non-bacterial prostatitis.