• The Journal of Korean Society of Virology
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• v.30 no.2
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• pp.125-130
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• 2000

Infection with human cytomegalovirus (HCMV) is of considerable clinical relevance after placental transmission and in immunosuppressed patients such as transplant recipients or patients with AIDS. The rapid detection method of HCMV has been required to overcome the time-consuming methods such as classical plaque assay or other immunological methods. This study was performed to establish the in situ ELISA, in which human lung fibroblasts infected with HCMV were fixed and used directly as antigen in 96 well culture plate. Expressed HCMV antigens were detected with HCMV-specific monoclonal antibodies. This method could detect HCMV dose-dependently upto $3{\times}10^2\;pfu/ml$. Antiviral activity of ganciclovir could be assayed within the known range of effective dose. This result showed that HCMV could be quantitated by in situ ELISA. The chemical, which was selected on the basis of component analysis in natural product, was tested to have the anti-HCMV activity by in situ ELISA, and three among five samples were found to have anti-HCMV activity with the dose-dependent manner. Conclusively in situ ELISA could be useful method for quantitation of HCMV and screening antiviral activity of samples to HCMV.

• The Microorganisms and Industry
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• v.18 no.2
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• pp.26-30
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• 1992

HIV는 ELISA나 WB에 의해 항체가 검출되기 수개월 혹은 수 년전에도 proviral DNA 상태로 감염된 세포의 chromosome내에 존재하는 것이 주지의 사실이다. 그동안 Southern blot, in situ hybridization등에 의해 이 proviral DNA를 검출하려는 연구가 진행되어 왔으나 lymphocyte $10^{4}$-$10^{6}$개 중 1개가 감염되어 있으며 lymphocyte chromosomal DNA에 비해 viral DNA의 양이 미량이므로 검출하기에는 민감도가 낮은 문제점이 있다. 본 고에서는 근래 개발되어 널리 사용되고 있는 polymerase chain reaction(PCR)을 이용한 HIV의 진단에 관해 살펴보고자 한다.

• Clinical and Experimental Reproductive Medicine
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• v.32 no.2
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• pp.101-111
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• 2005

배 경: Pituitary adenylate cyclase-activating polypeptide (PACAP)은 양의 시상하부에서 추출된 신경펩타이드 호르몬으로 난소에도 존재하여 배양된 과립막 세포에서 스테로이드합성과 cyclic AMP 형성을 촉진함이 보고되었다. 목 적: 흰쥐 난소를 실험 모델로 사용하여 배란시 황체화호르몬 (luteinizing hormone; LH)에 의해 유도된 PACAP과 PACAP 수용체의 유전자 발현양상과 신호 전달경로를 규명하고자 하였다. 재료 및 방법: 미성숙 흰쥐의 배란전 난포를 체외 배양하면서 LH로 처리하고 PACAP 및 PACAP수용체의 유전자 발현을 보기 위해서는 Northern blot 분석과 in situ hybridization (ISH)을, 그리고 단백질 수준의 PACAP 검색을 위해서는 enzyme linked immunosorbent assay (ELISA) 분석을 이용하였다. 결 과: LH 처리 후 Northern blot상의 PACAP 유전자 발현은 6~9시간에 일시적으로 최고치에 도달하였으며 ISH로 보아 과립막 세포에서 발현됨을 알 수 있었다. ELISA 분석 상 PACAP 단백질도 LH처리 후 6~12시간에 최고치를 나타내었으며, PACAP 수용체 mRNA 역시 3~9시간에 최고치로 과립막 세포에서 발현되었다. Adenylate cyclase (AC) 억제제인 MDL12330A 처리시 LH로 발현된 PACAP mRNA가 감소되며, AC의 활성제인 forskolin 처리에는 LH시와 유사한 PACAP mRNA의 발현양상을 나타내었다. 그러나 protein kinase C (PKC)의 억제제인 chelerythrine과 2-0-tetradecanolphorbol-13-acetate (TPA) 처리로는 PACAP 의 유전자 발현에 영향을 주지 못하였다. 5-lipoxygenase의 억제제인 MK886이나 nordihydroguaiaretic acid (NDGA)로 처리한 결과 LH로 유도된 PACAP 유전자의 발현이 감소되었으나, cyclooxygenase의 억제제인 indomethacin은 별로 영향을 주지 못하였다. MEK와 p38의 억제제인 PD98059와 SB203580도 LH로 촉진 된 PACAP의 유전자 발현을 농도 의존적으로 억제하였다. 결 론 : 배란전 난포에서 PACAP과 PACAP 수용체의 유전자 발현은 모두 LH의 폭발적 분비에 의해 유도되어 일시적으로 과립막 세포에서 나타나 배란을 위한 국소적인 조절 작용을 할 것으로 추정되며, LH로 촉진된 PACAP 유전자 발현을 위한 신호전달은 cAMP-PKA, lipoxygenase 및 MAP kinase 경로를 통하는 것으로 사료된다.

• Asian Pacific Journal of Cancer Prevention
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• v.13 no.10
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• pp.5027-5031
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• 2012

This study was conducted to investigate the expression of survivin and caspase 3 in oral squamous cell carcinoma and peritumoral tissue, and possible pathogenesis mechanisms. We used ELISA and western blotting to detect the protein expression levels of survivin and caspase 3 in tissue. In situ hybridization and real-time PCR were applied to assess mRNA expression levels. In this study, 13 tumor samples and 13 peritumoral tissue samples were collected from oral squamous cell carcinoma patients and 10 normal tissue samples obtained from patients without tumor. The result showed that the protein and mRNA expression of survivin in carcinoma was the highest among three types of tissue; following was that in peritumoral tissue. No difference in caspase 3 zymogen between peritumoral tissue and normal tissue could be found, while it was evidently decreased in carcinoma tissue. Activated caspase 3 was detected in normal tissue but could not be identified in peritumoral or carcinoma tissue. Our results indicate that the expression of survivin is apparently elevated in tumoral and peritumoral tissue. Expression of activated caspase 3 was not detected in tumoral tissue and the expression of caspase 3 zymogen was decreased in tumoral tissue. Our findings suggest that survivin may inhibit both synthesis and activation of caspase 3, hence inhibiting cell apoptosis and facililitating eventual development of oral squamous cell carcinoma.

• Journal of Aquaculture
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• v.15 no.1
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• pp.7-13
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• 2002

Since 1993, the White Spot Syndrome Virus (WSSV) disease occurred in China among cultured shrimps resulting in mass mortality. Epizootiological surveys undertaken during the outbreak period of 1993-1994 indicated that all stages of Penaeus chinensis, P. japonicus and P. monodon were infected. Consequent to the transport of contaminated shrimp seedlings and seawater, the disease spread all over the farms of China. The disease was more rapidly transmitted at temperatures above $25^{\circ}C$. Challenge experiments showed the causative agent was highly virulent. White spots appeared on the carapace of both span-taneous and experimentally infected shrimps. Moribund shrimps contained turbid hemolymph, hypertrophied Iymphoid organ and a necrotic mid-gut gland. Electron microscopy showed the presence of viral particles in the gills, stomach, lymphoid organ, and epidermal tissue of the infected shrimp. The visions were slightly ovoid with an envelope and averaged 350 $\times$ 150 nm; nucleocapsids measured 375 $\times$ 157 nm. With discontinuous sucrose gradient of 35, 50 and 60% (w/v), the virus was separated from hemolymph of the infected shrimp. The estimated molecular weight of genomic DNA was 237 Kb with EcoR I, 247 Kb with Hind III and 241kb with Pst I. A total of 9 hybridoma colones secreting monoclonal antibodies (MAbs) were produced from mouse myeloma and spleen cells immunized with WSSV. The immunofluorescence assay of gill tissue showed that the MAbs reacted with diseased but not with healthy shrimp. The MAbs belonged to IgGl, IgG2b subclass and IgM class, all with kappa light Immune-electron-microscopy with colloidal gold marker showed the presence of 5 MAbs epitopes on the envelope and one on the capsid of the virus. Baculoviral mid-gut gland necrosis showed the specificity of the MAbs produced. For diagnosis 5 different methods were selected. Using Kimura primers for PCR, or MAbs for immunoblot, ELISA or FAT method, in situ hybridization was carried out to show the gene. All these methods detected WSSV in the organ samples of the diseased shrimp but not in healthy one.