• Title/Summary/Keyword: ELISA method

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Tumor Inhibition Effects and Mechanisms of Angelica sinensis and Sophorae flavescentis ait Decoction Combined with Cisplatin in Xenograft Mice

  • Yan, De-Qi;Liu, Yong-Qi;Li, Ying-Dong;Li, Dou;Cheng, Xiao-Li;Wu, Zhi-Wei
    • Asian Pacific Journal of Cancer Prevention
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    • v.15 no.11
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    • pp.4609-4615
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    • 2014
  • Background: To investigate tumor inhibition effects and mechanisms of Angelica sinensis and Sophorae flavescentis ait decoction (ASSF) combined with diamine-dichloroplatinum (DDP). Materials and Methods: Bodyweight, tumor inhibition rate and q value were calculated for single ASSF or ASSF combined with DDP on H22 carcinoma xenograft KM mice. Biochemical methods for serum LDH, AST, ALT, and AKP, ELISA method for serum HIF-$1{\alpha}$, pathological assessemnt of thymus, immunohistochemistry detection of tumor tissue caspase3 and mutant p53 protein, and qRT-PCR detection of bax/ bcl-2 mRNA were applied. Results: Compared with DDP control group, the bodyweight increased in ASSF-DDP group (p<0.01). Tumor inhibition rates for DDP, ASSF, ASSF-DDP were 62.7%. 43.7% and 71.0% respectively, with a q value of 0.90. Compared with other groups, thymus of DDP control group had obvious pathological injury (p<0.01), serum LDH, AST, ALT, AKP increased significantly in DDP control group (p<0.01), while serum HIF-$1{\alpha}$ was increased in the model control group. Compared with this latter, the expression of mutant p53 protein and bcl-2 mRNA were decreased in all treatment groups (p<0.01), but there were no statistical difference between DDP control p and ASSF-DDP groups. The expression of caspase3 protein and bax mRNA was increased in all treatment groups, with statistical differences between the DDP and ASSF-DDP groups (p<0.01). Conclusions: ASSF can inhibit bodyweight decrease caused by DDP, can inhibit tumor growth synergistically with DDP mainly through increasing serum HIF-$1{\alpha}$ and pro-apoptotic molecules such as caspase 3 and bax, rather than through decreasing anti-apoptotic mutant p53 and bcl-2. ASSF can reduce DDP toxicity due to decreasing the release of LDH, AST, ALT, AKP into blood and enhancing thymus protection.

Changes in Antioxidant Activity during Growth of Artemisia iwayomogi (더위지기 생육 중 항산화 활성 변화)

  • Hwang, Tay-Eak
    • Korean Journal of Medicinal Crop Science
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    • v.17 no.4
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    • pp.286-292
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    • 2009
  • The aim of this study was to investigate the effect of plant growth at several different growing periods on antioxidant activities and zeatin and ABA contents of Artemisia iwayomogi. Measurements of antioxidant activities, lipid peroxidation inhibition, and superoxide radical scavenging activity were done using PMS, NBT and lipid auto-oxidation analysis, respectively. The results show that activities of antioxidants from Artemisia iwayomogi had much higher than BHT. DPPH free radical scavenging effect of Artemisia leaf extract was increased from $71.06{\pm}4.36%$ in April to $90.06{\pm}4.41%$ in October. Activities of superoxide radical scavenging and lipid peroxidation inhibition were $33.83{\pm}3.45%$ and $45.60{\pm}3.10$ in April and then increased to $84.40{\pm}4.00%$ and $75.86{\pm}3.50%$ in October, respectively. An ELISA technique has been developed for the determination of zeatin and ABA in Artemisia leaves. By this method, the content changes of zeatin and ABA from Artemisia during the growth were investigated. The zeatin content in leaf was measured to be $186.86{\pm}1.40$ pmol/g dry weight in April, however, decreased to $117.93{\pm}5.83$ pmol in October. The ABA content in leaf increased from $19.00{\pm}1.26$ nmol in April to $68.20{\pm}2.52$ nmol in October. Relationship between antioxidant activities and plant hormone contents was indicated that antioxidant activity may depend on decreasing zeatin content or increasing ABA content.

Effect of the low level light irradiation to NTacSam:SD tissue cell culture (NTacSam:SD의 조직세포 배양에 저출력 광원의 효과)

  • Kim, Tae-Gon;Kim, Toung-Pyo;Park, No-Bong;Lee, Ho-Sic;Park, Yong-Pil;Cheon, Min-Woo
    • Proceedings of the Korean Institute of Electrical and Electronic Material Engineers Conference
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    • 2009.06a
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    • pp.423-423
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    • 2009
  • Currently, lasers are one of the most popular light sources in use for medical treatment. Many studies on low power lasers are being done in cell culture or through animal tests and most report different findings, making it difficult to verify their true effects. There are shifts in trends of studies from laser and LED that are expensive and generate heat problem to LED that are economically effective and safe. Its near infrared rays can penetrate deep into skin or muscle, up to 23 cm, without causing thermal damage or impairing neighboring tissues. This study verified the performance and effectiveness of an LED irradiator that was designed to emit similar wavelengths to that of a laser and thus could be used instead of a low level laser therapy in experiments on animals. And then, each experiment was performed to irradiation group and non-irradiation group for NTacSam:SD tissue cells. MIT assay method was chosen to verify the cell increase of two groups and the effect of irradiation on cell proliferation was examined by measuring 590nm transmittance of ELISA reader. As a result, the cell increase of NTacSam:SD tissue cells was verified in irradiation group as compared to non-irradiation group. The fact that specific wavelength irradiation has an effect on cell vitality and proliferation is known through this study.

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Characteristics of Low-level Light Source for Animal Cell Proliferation (동물 세포 증식을 위한 저출력 광 소스의 특성)

  • Cheon, Min-Woo;Kim, Seong-Hwan;Song, Chang-Hun;Mun, Seong-Pyo;Kim, Tae-Gon;Park, Yong-Pil;Kim, Dae-Young;Kim, Young-Su
    • Journal of the Korean Institute of Electrical and Electronic Material Engineers
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    • v.20 no.1
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    • pp.92-97
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    • 2007
  • This paper performed the basic study for developing the Photodynamic Therapy Equipment for medical treatment. We developed the equipment palpating cell proliferation using a high brightness LED. This equipment was fabricated using a micro-controller and a high brightness LED, and designed to enable us to control light irradiation time, intensity, frequency and so on. Especially, to control the light irradiation frequency, FPGA was used, and to control the change of output value, TLC5941 was used. Control stage is divided into 30 levels by program. Consequently, the current value could be controlled by the change of level in Continue Wave(CW) and Pulse Width Modulation(PWM), and the output of a high brightness LED could be controlled stage by stage. And then, each experiment was performed to irradiation group and non-irradiation group for both Rat bone marrow and Rat tissue cells. MTT assay method was chosen to verify the cell increase of two groups and the effect of irradiation on cell proliferation was examined by measuring 590 nm transmittance of ELISA reader. As a result, the cell increase of Rat bone marrow and tissue cells was verified in irradiation group as compared to non-irradiation group. The fact that specific wavelength irradiation has an effect on cell vitality and proliferation is known through this study.

Apoptotic Process is Involved in the L-Glutamate-Induced PC12 Cell Death (L-Glutamate에 의한 PC12 세포의 고사성 사망)

  • Sung, Ki-Wug;Jung, Kyung-Heui;Kim, Seong-Yun;Kang, Jung-Hyae;Lee, Sang-Bok
    • The Korean Journal of Physiology and Pharmacology
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    • v.1 no.6
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    • pp.699-705
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    • 1997
  • Although it is known that neuronal cell death during development occurs by apoptosis, the mechanisms underlying excitatory amino acid-induced neuronal cell death remain poorly understood. In this study we have examined the mechanism by which L-glutamate, an excitatory amino acid neurotransmitter, induces cell death in PC12 cell lines. To characterize cell death, we employed sandwich enzyme-linked immunosorbent assay(ELISA) method for cellular DNA fragmentation, DNA agarose gel electrophoresis and chromatin staining by acridine orange and ethidium bromide after treating the PC12 cells with L-glutamate. L-Glutamate caused dose-dependent cell death with a maximum at 24 hrs after the treatment. These cellular fragmentation was blocked by pretreatment of MK-801, a noncompetitive N-methyl-D-aspartic acid(NMDA) receptor antagonist, and nerve growth factor(NGF). Analysis of DNA integrity from L-glutamate-treated cells revealed cleavage of DNA into regular sized fragments, a biochemical hallmark of apoptosis. The PC12 cells that were induced to die by L-glutamate treatment exhibited classical chromatin condensation under the light microscopy after acridine orange and ethidium bromide staining. These results suggest that apoptosis is one of the key features that are involved in L-glutamate-induced excitotoxic cell death in PC12 cells, and these cell death are mediated by NMDA receptor and depend on NGF.

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Development of Radioimmunoassay(RIA) for Residue Analysis with Endosulfan in Water and Carp(Cyprinus carpio L.) (잉어중 Endosulfan의 잔류분석(殘留分析)을 위한 Radioimmunoassay(RIA)의 개발(開發))

  • Lee, Kang-Bong;Shim, Jae-Han;Suh, Yong-Tack
    • Korean Journal of Environmental Agriculture
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    • v.13 no.3
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    • pp.262-270
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    • 1994
  • The established methods in the residue analysis of endosulfan require an extensive sample clean-up prior to quantification by relatively complex equipment. A radioimmunoassay(RIA) provides a simple procedure with theoretically higher sensitivity and specificity necessitating only a minimum of sample clean-up. Endosulfan-specific antibodies were developed in rabbits by using a bovine serum albumin(BSA) conjugate wherein the alcohol form of endosulfan was multiply bound to the protein via succinylation. Produced antibodies showed the high titers to endosulfan-BSA(1 : 32,000). An RIA method was developed in water and carp by using $^{14}C-labeled$ endosulfan as a tracer. The lowest detection amount of endosulfan was 1 ng in the liver, kidneys, gut and water samples, and 3 ng in the whole body sample of carp without any clean-up, corresponding to 0.1 ppb of endosulfan.

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Electrochemical Immunosensor Based on the ZnO Nanorods Inside PDMS Channel for H7N9 Influenza Virus Detection (PDMS 채널 내부에 성장된 산화아연 나노막대를 이용한 H7N9 인플루엔자 바이러스 전기화학 면역센서)

  • Han, Ji-Hoon;Lee, Dongyoung;Pak, James Jungho
    • Journal of Sensor Science and Technology
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    • v.23 no.4
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    • pp.278-283
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    • 2014
  • In this study, we propose an immunosensor using zinc oxide nanorods (NRs) inside PDMS channel for detecting the influenza A virus subtype H7N9. ZnO with high isoelectric point (IEP, ~9.5) makes it suitable for immobilizing proteins with low IEP. In this proposed H7N9 immunosensor structure ZnO NRs were grown on the PDMS channel inner surface to immobilize H7N9 capture antibody. A sandwich enzyme-linked immunosorbent assay (ELISA) method with was used 3,3',5,5' tetramethylbenzidine (TMB) for detecting H7N9 influenza virus. The immunosensor was evaluated by amperometry at various H7N9 influenza antigen concentrations (1 pg/ml - 1 ng/ml). The redox peak voltage and current were measured by amperometry with ZnO NWs and without ZnO NWs inside PDMS channel. The measurement results of the H7N9 immunosensor showed that oxidation peak current of TMB at 0.25 V logarithmically increased from 2.3 to 3.8 uA as the H7N9 influenza antigen concentration changed from 1 pg/ml to 1 ng/ml. And then we demonstrated that ZnO NRs inside PDMS channel can improve the sensitivity of immunosensor to compare non-ZnO NRs inside PDMS channel.

Development of Rapid, Safe Analytical Techniques of Aflatoxins and Their Current Regulation (Aflacxin에 대한 최신 분석법과 규제동향)

  • 정덕화
    • Journal of Food Hygiene and Safety
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    • v.5 no.3
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    • pp.131-138
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    • 1990
  • Aflatoxins is a chemically diverse group of toxic secondary metabolites that are produced by fungi and often occur in agricultural commodities. Because of their wide range of toxic effects, Aflatoxins cause severe economic losses to farmers and livestock producers and pose a health to human consuming contaminated foods. Long term prospects for biotechnological control of Aflatoxins require elucidation of the specific steps and regulation of their biosynthetic pathways . Aflatoxin determinations can be approached many ways. It is essential to safely handle all experimental materials associated with aflatoxin analysis or aflatoxigenic fungi Visual screening of suspect samples, base on the presence of conidial head of the aspergillus flavus group, and screening samples for the presence of bright greenish yellow flourescence are not chemical tests and such screening techniques may allow aflactoxin contaminated lots into commerce. Microcolumn screening procedures should always be used in conjunction with a quantitative method. Several thin layer chromatography(TLC) and high performance liquid chromatography(HPLC) methods are suitable for quantitation and are in general use. Immunochemical Methods such as the ELISA or affinity column chromatography methods are being rapidly developed. The chemical and immunochemical methods can be reliable if care is taken, using suitable controls and personnel that are well trained . All analytical laboratories should stress safety and include suitable analytical validation procedure. Especially a worldwide enquiry was undertaken in recent to obtain up-to-date information about aflatoxin legislation in as many countries of the world as possible. The information concerns aflatoxin in foodstuffs. aflatoxin MI in dairy products, aflatoxins in animal feedstuffs. Limits and regulations for aflatoxin have been expended in recent with more countries having legislation on subject, more products, and more aflatoxins covered by this legislation.

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The Effects of Bee Venom on PLA2, COX-2, iNOS, AA and PG in RAW 264.7 Cells (봉약침액(蜂藥鍼液)이 PLA2, COX-2, iNOS, AA 및 미치는 영향(影響)에 관(關)한 연구(硏究))

  • Ha, Seang-Jong;Lee, Seong-No;Jo, Hyun-Chul;Kim, Kee-Hyun
    • Journal of Pharmacopuncture
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    • v.5 no.2
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    • pp.40-51
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    • 2002
  • Objectives : The purpose of this study was to investigate the effect of Bee Venom on the lipopolysaccharide-induced expression phospholipase $A_2$, cyclooxygenase-2 and inducible nitrogen oxide synthase, and the generation of arachidonic acid, prostaglandin D2 and E2 in RAW 264.7 cells, a murine macrophage cell line. Methods : The expression of phospholipase $A_2$, cyclooxygenase and inducible nitrogen oxide synthase was determined by western blotting with corresponding antibodies, and the generation of arachidonic acid, prostaglandin $D_2$ and $E_2$ was assayed by ELISA method in RAW 264.7 cells. The non-toxic concentrations (0.1 to $5\;{\mu}g/ml$) of bee venom determined by MTT assay, were used in this study. Results : 1. Bee venom inhibited lipopolysaccharide-induced expression of phospholipase $A_2$ in a dose dependent manner after 48 hours treatment. 2. Bee venom inhibited lipopolysaccharide-induced expression of cyclooxygenase-2 in a dose dependent manner after 24 and 48 hours treatment. 3. Bee venom inhibited lipopolysaccharide-induced expression of inducible nitrogen oxidesynthase in a dose dependent manner after 48 hours treatment. 4. The generation of arachidonic acid, prostaglandin $D_2$ and $E_2$ was not much affected by the treatment of bee venom on the lipopolysaccharide-induced generation of arachidonic acid, prostaglandin $D_2$ and $E_2$ in RAW 264.7 cells.

Development of a Rapid Diagnostic Test Kit to Detect IgG/IgM Antibody against Zika Virus Using Monoclonal Antibodies to the Envelope and Non-structural Protein 1 of the Virus

  • Kim, Yeong Hoon;Lee, Jihoo;Kim, Young-Eun;Chong, Chom-Kyu;Pinchemel, Yanaihara;Reisdorfer, Francis;Coelho, Joyce Brito;Dias, Ronaldo Ferreira;Bae, Pan Kee;Gusmao, Zuinara Pereira Maia;Ahn, Hye-Jin;Nam, Ho-Woo
    • Parasites, Hosts and Diseases
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    • v.56 no.1
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    • pp.61-70
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    • 2018
  • We developed a Rapid Diagnostic Test (RDT) kit for detecting IgG/IgM antibodies against Zika virus (ZIKV) using monoclonal antibodies to the envelope (E) and non-structural protein 1 (NS1) of ZIKV. These proteins were produced using baculovirus expression vector with Sf9 cells. Monoclonal antibodies J2G7 to NS1 and J5E1 to E protein were selected and conjugated with colloidal gold to produce the Zika IgG/IgM RDT kit (Zika RDT). Comparisons with ELISA, plaque reduction neutralization test (PRNT), and PCR were done to investigate the analytical sensitivity of Zika RDT, which resulted in 100% identical results. Sensitivity and specificity of Zika RDT in a field test was determined using positive and negative samples from Brazil and Korea. The diagnostic accuracy of Zika RDT was fairly high; sensitivity and specificity for IgG was 99.0 and 99.3%, respectively, while for IgM it was 96.7 and 98.7%, respectively. Cross reaction with dengue virus was evaluated using anti-Dengue Mixed Titer Performance Panel (PVD201), in which the Zika RDT showed cross-reactions with DENV in 16.7% and 5.6% in IgG and IgM, respectively. Cross reactions were not observed with West Nile, yellow fever, and hepatitis C virus infected sera. Zika RDT kit is very simple to use, rapid to assay, and very sensitive, and highly specific. Therefore, it would serve as a choice of method for point-of-care diagnosis and large scale surveys of ZIKV infection under clinical or field conditions worldwide in endemic areas.