• 한국동물위생학회지
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• 제46권2호
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• pp.123-135
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• 2023

Feline calicivirus (FCV) is considered the main viral pathogen of feline upper respiratory tract disease (URTD). The frequent mutations of field FCV strains result in the poor diagnostic sensitivity of previously developed molecular diagnostic assays. In this study, a more sensitive real-time reverse transcription-polymerase chain reaction (qRT-PCR) assay was developed for broad detection of currently circulating FCVs and comparatively evaluated the diagnostic performance with previously developed qRT-PCR assay using clinical samples collected from Korean cat populations. The developed qRT-PCR assay specifically amplified the FCV p30 gene with a detection limit of below 10 copies/reaction. The assay showed high repeatability and reproducibility, with coefficients of intra-assay and inter-assay variation of less than 2%. Based on the clinical evaluation using 94 clinical samples obtained from URTD-suspected cats, the detection rate of FCV by the developed qRT-PCR assay was 47.9%, which was higher than that of the previous qRT-PCR assay (43.6%). The prevalence of FCV determined by the new qRT-PCR assay in this study was much higher than those of previous Korean studies determined by conventional RT-PCR assays. Due to the high sensitivity, specificity, and accuracy, the new qRT-PCR assay developed in this study will serve as a promising tool for etiological and epidemiological studies of FCV circulating in Korea. Furthermore, the prevalence data obtained in this study will contribute to expanding knowledge about the epidemiology of FCV in Korea.

• Toxicological Research
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• 제21권2호
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• pp.175-178
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• 2005

Butyl $\rho$-hydroxybenzoic acid (butyl paraben, BP) is a homologous series of parabens and is widely used as a preservative in cosmetic and pharmaceutical products. The purpose of this study was to investigate the estrogenic/antiandrogenic activities of BP in animals. For that, we performed an uterotrophic assay and a Hershberger assay in rats. In uterotrophic assay, BP was administered subcutaneously to immature female SD rats (18 days old) for 3 consecutive days. The wet and dry uterus weights were significantly increased in the groups treated with BP in dose­dependent manner. In case of Hershberger assay, BP significantly reduced the weight of seminal vesicle of castrated rats. And other accessory organ/glands - prostate, Cowper's glands, bulbocavernosus muscle and glans penis were also slightly decreased. The results of this study suggested that BP showed estrogenic and anti-androgenic activities in vivo.

• 한국환경성돌연변이발암원학회지
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• 제17권2호
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• pp.71-77
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• 1997

The single cell gel electrophoressis(SCGE) assay, also known as the comet assay, is a rapid, simple, visual and sensitive technique for measuring and analysing DNA breakage in mammalian cells. The SCGE or comet assay is a promising test for the detection of DNA damage and repair in individnal cells. It has widespread potential applications in DNA damage and repair studies, genotoxicity testing and biomonitoring. In this microgel electrophoresis technique, cells are embedded in agarose gel on microscope slides, iysed and electrophoresed under alkaline conditions. Cells with increased DNA damage display increased migration of DNA from the nucleus towards the anode. The length of DNA migration indicates the amount of DNA breakage in the cell. The comet assay is also capable of identifying apoptotic cells which contain highly fragmented DNA. Here we review the development of the SCGE assay, existing protocols for the detection and analysis of comets, the relevant underlying principles determining the behaviour of DNA and the potential applications of the technique.

• 한국환경성돌연변이발암원학회지
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• 제18권2호
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• pp.129-134
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• 1998

To evaluate the genotoxicity of CKD-602, an anticancer agent the in viかo reverse mutation assay using Salmonella typhimurium, the Chromosome aberration assay using Chinese hamster lung (CHL) cells and the in vivo micronucleus assay using bone marrow cells of ddY mice were performed. In the reverse mutation assay, CKD-602 did not induced mutagenicity in Salmonella typhimurium TA 98, TA 100, TA 1535, and TA 1537 strains with and without metabolic activation. In the chromosome aberration test using CHL cells, there was an increased incidence of structural aberrations induced by CKD-602 without metabolic activation during 24 and 48 hours, but CKD-602 did not induce chromosome aberration with metabolic activation. The in vivo induction of micronuclei was measured in polychromatic erythrocytes of bone marrow of male ddY mice. At 24 hours after treatment with CED-602 by i.p. once, there was an increased incidence of micronucleated polychromatic erythrocytes in bone marrow of ddY male mice.

• 한국환경성돌연변이발암원학회지
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• 제18권2호
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• pp.123-128
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• 1998

To evaluate the genotoxicity of SKP-450, an antihypertensive agent the in vitro reverse mutation assay using Salmonella typhimurium, the Chromosome aberration assay using Chinese hamster lung (CHL) cells and the in vivo micronucleus assay using bone marrow cells of ddY mice were performed. In the Reverse mutation test, SKP-450 did not induced mutagenicity in Salmonella typhimurium TA 98, TA 100, TA 1535 and TA 1537 with and without metabolic activation. In the chromosome aberration assay using CHL cells, there was no increased incidence of structural and numerical aberrations with and without metabolic activation. The in vivo induction of micronuclei was measured in polychromatic erythrocytes of bone marrow of male ddY mice at 30 hours after treatment with SKP-450 by p.o once. The results showed no increased incidence of micronucleated polychromatic erythrocytes in bone marrow of ddY male mice treated with SKP-450.

• 한국임상수의학회지
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• 제13권2호
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• pp.158-162
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• 1996

Comet assay 는 포유류 세포에서 DNA의 파괴를 측정하는데 있어 신속하고 단순하며 시각적이고 민감한 방법이다. Apoptosis에서는 세포핵의 광범위한 DNA의 붕괴가 일어나므로 comet assay는 종양세포에서 apoptosis가 발생되었는가를 알아내는데 유용하다. 본 연구는 apoptosis 연구의 결과가 변화되지 않도록 comet assay slides를 좀 더 오래 보관할 수 있는 방법을 개발하고자 시행되었다. 개의 종양세포를 가지고 alkali comet assay를 끝낸 뒤 slides를 진공 건조기에서 꺼내서 증류수로 점적하여 10-20분간 침수시키고 현광현미경하에서 육안적으로 관찰하였다. 건조후 3-4일, 1주, 2주, 3주, 4주 및 7주의 slides에서 apoptosis 회복율(%)은 각각 98.1, 98.3, 99.4, 80.8 및 35.2%이었다. 3주 이내의 slides에서 대조군과 비교하여 apoptosis 회복율에서 차이가 없었으나 4주 이상의 slides를 건조후 침수시키는 방법을 이용하였을 때 apoptosis 평가에서 건조 후 3주간까지는 처음의 결과와 차이가 없으며, 이 방법을 이용하여 comet slide의 좀 더 긴 기간이 보관과 보관후의 재평가에서 이용될 수 있는 좋은 방법이 된다.

• 한국환경성돌연변이발암원학회지
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• 제21권1호
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• pp.44-50
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• 2001

To identify nongenotoxic carcinogen determined as negative by ICH guideline-recommended standard genotoxicity test battery; Ames test, chromosome aberration assay, mouse lymphoma $tk^{+/-}$ assay, in vivo micronucleus assay, we picked bisphenol A as a model compound. In this study, we applied in vitro BalB/c 3T3 cell transformation assay and Syrian hamster embryo (SHE) cell transfarmation assay. Bisphenol A was treated upto $769.2 ug/m{\ell}$ in BalB/c 3T3 cells and upto $125 ug/m{\ell}$ in SHE cells. bisphenol A didn't induced morphological transformation both with one stage treatment protocol and with two stage treatment protocol. But, treated far 48 hr, Bisphenol A induced morphological transformation significantly in SHE cells.

• 자연과학논문집
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• 제9권1호
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• pp.25-29
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• 1997

Glucocorticoid에 의한 항염증 작용의 second messenger로 생각되어지는 annexin superfamily중 하나인 37 kDa의 단백질, lipocortin-1의 작용기작을 이해할 목적으로 in vivo에서 protein-protein interaction을 인식하는 yeast-based genetic assay인 yeast two assay를 통하여 lipocortin-1과 결합하는 단백질 유전자를 분리하여 조사하였다. 이 방법으로 실험을 수행한 결과 분리된 유전자가 human serine proteinase 유전자와 homology가 있는 것으로 밝혀졌다.

• Bulletin of the Korean Chemical Society
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• 제22권4호
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• pp.362-366
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• 2001

To show the effects of metallothionein (MT) on the activity of enzymes involved in the removal of reactive oxygen species, MT has been added to the assay systems of superoxide dismutase (SOD), catalase and peroxidase. We have used assay systems of SOD based on NADPH oxidation and nitrite formation from hydroxylammonium chloride as an assay of superoxide breakdown rate. The two assay systems showed different results at the high concentration of MT. MT showed the scavenging of superoxide in the SOD assay system in the presence and absence of SOD. MT added to the SOD assay system behaved as an activator of SOD, but apo-MT behaved as an inhibitor. When MT was added to the assay system in the presence of a fixed amount of SOD, the breakdown rate of superoxide increased. The effects of MT on the decomposition of hydrogen peroxide and the activity of catalase and peroxidase decomposing hydrogen peroxide were evaluated. MT decreased the activities of catalase and peroxidase. We have concluded that the function of MT as an antioxidant might effect the level of superoxide scavenging and not the level of hydrogen peroxide.

• Radiation Oncology Journal
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• 제19권2호
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• pp.163-170
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• 2001

목적 : 방사선조사후 세포의 생존 분획은 세포집락 측정기법으로 확인하는 것이 표준이나 많은 비용과 시간이 소요되는 단점을 갖고 있다. 이에 생존 세포의 tetrazolium염의 자색 formazan 침전물로의 환원시키는 능력에 그 기반을 둔 MTT 기법을 사용하여 세포집락 측정기법을 대체하기 위한 기법으로서의 유용성과 그 실행상의 최적조건을 규명하고자 하였다. 방법 : PCI-1, SNU-1066, NCI-H630, RKO등의 세포주에 0, 2, 4, 6, 8, 10 Gy의 방사선을 조사한 후 세포집락 측정 기법과 MTT기법으로 세포 생존 분획을 조사하였다. 세포집락 측정기법은 $25\;cm^2$ 폴리스티렌 배양 플라스크에 방사선량에 따라 다른 수의 세포를 분주한 후 24시간 동안 배양 후 방사선을 조사하였고 이를 $10\~14$일 동안 배양 후 염색하여 생성된 세포집락의 수를 측정하였다. MTT기법은 침전물의 용해과정이 필요없는 Premix WST-1 시약을 이용하여 시행하였다. MTT기법은 각각의 세포주에서 세포수와 흡광도간의 선형관계와 최적 실험조건을 확인한 이후 시행하였다. 이 기법은 방사선을 조사받은 세포에서는 지수적 성장을 회복한 이후와 방사선을 조사받지 않은 세포는 4회 이상의 세포분열을 거친 후에 시행하였다. 세포집락 측정기법 및 MTT기법을 통하여 얻은 세포 생존율을 구한 후 이를 지표로 비교하였다. 결과 : 각 기법으로 얻은 세포 생존율의 표준편차는 $5\%$ 내외였다. 2가지 방법으로 구한 세포 생존율은 t-test로 비교하였을 때 $0\~4\;Gy$에서는 통계적으로 유의한 차이가 없었으며 회귀분석 결과는 선형적 관계가 있었다$(R^2=0.975-0.992)$. MTT 기법의 시행에 최적인 세포수는 배양효율에 따라 다른 것으로 나타났는데, 배양효율이 $30\%$ 이상이면 300개 이하가, $30\%$ 미만인 경우는 $500\~1,000$개가 적당한 것으로 확인되었다. MTT기법은 6 배가시간 경과 이후에 시행하는 것이 세포집락 측정기법과 가장 근접하였으며 적어도 4 배가시간 이후에 시행하는 것이 필요할 것으로 사료되었다. 이에 따르면 배가시간이 3일 이하인 세포주가 세포 민감성 측정방법으로서 MTT 기법이 세포 집락 측정 기법을 대체하여 사용하기에 적합한 것으로 사료되었다. 결론 : 이상에서, MTT기법을 이용하여 방사선조사 후의 세포생존을 측정하기 위해서는 예비실험을 통해 각 세포주에서의 최적의 조건을 찾는 것이 필수적이며 이 조건하에서 MTT기법을 시행해야만 방사선에 의한 세포 민감성 측정에 이용될 수 있음을 확인하였다.