• 한국동물학회지
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• 제15권3호
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• pp.101-104
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• 1972

간편하고 경제적이며 재현성도 우수한 cellulose acetate 전기영동장치를 제작하였다. 본 장치를 사용하여 몇가지 동물 조직의 lactate 및 malate dehydrogenase isozyme 을 분리 검출하여 본 결과, 기존 제품으로 실험한 결과와 별로 차이가 없음을 알았다. 흔히 cellulose acetate 전기영동은 일정한 정전류에서 실시하는데, 본 장치에서는 약간의 전류변동이 있는 것이 사실이나 실험결과는 충분히 실용적인 것이었다.

• 동의생리병리학회지
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• 제18권3호
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• pp.754-758
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• 2004

To test the cytoprotective effect of sophorae radix (SR) against hydrogen peroxide (H₂O₂)-induced cytotoxicity, we investigated the cell viability using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay in the presence of methylene chloride, n-butanol, ethyl acetate and water soluble fraction of SR water extracts in H9c2 cells. These results were obtained as followed; H₂O₂ decreased the cell viability of H9c2 cells in a dose dependent manner. Cells pretreated with SR water extracts were protected the H₂O₂-induced decrease of viability in H9c2 cells. Among organic solvents fractions of SR water extracts, ethyl acetate soluble fractions of SR protected the decrease of viability induced by H₂O₂ in H9c2 cells. These results suggest that ethyl acetate soluble fractions of SR water extracts is effective in the prevention of H₂O₂-induced cytotoxicity.

• Asian-Australasian Journal of Animal Sciences
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• 제21권4호
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• pp.494-498
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• 2008

MTT (3-(4, 5-dimethyl thiazol-2-yl)-2, 5-diphenyl tetrazolium bromide) reduction assay is a method that validates the viability of an active cell. Dehydrogenase in mitochondria converts yellow colored insoluble tetrazolium salt to purple colored water-soluble formazan. Sperm also have mitochondria in the midpiece, therefore sperm viability could be evaluated by MTT reduction assay. Several studies have already demonstrated the capability of application of the MTT reduction assay to sperm of several species in Hepes-BSA buffer. Because most liquid semen was diluted in extender like BTS (Beltsville Thawing Solution), Modena or Androhep when it is used or transferred, semen needed another dilution in Hepes-BSA buffer to assess sperm viability. In this study, we evaluated boar sperm viability especially in BTS extended semen and compared the efficiency of this test with eosin-nigrosin staining. We used the fresh BTS extended semen from a local A.I center. Semen sample was diluted to $3.0{\times}10^7$ sperms/ml in BTS. The rates of formazan production were measured in 96-well microtiter plates immediately and 1h after incubation at $17^{\circ}C$ using a spectrophotometer at wave length 560 nm. Simultaneously, split samples of the same semen were tested, using eosin-nigrosin staining to compare the efficiency of the MTT assay of sperm viability in BTS. The correlation between the results of these tests was calculated using Student-t test and ANOVA. The results revealed a strong correlation between the results of MTT reduction rate and the results that were simultaneously determined by eosin-nigrosin staining at 1 h. In conclusion, the MTT reduction test was an effective and simple method to validate sperm viability and it could be used as a simple tool to evaluate sperm viability in the local A.I center and laboratory.

• Toxicological Research
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• 제14권3호
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• pp.315-320
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• 1998

In vitro skin iritation of anti-inflammatory drugs was investigated in terms of the cytotoxicity method to human skin fibroblast cells. Five anti-inflammatory drugs (Diclofenac, Naproxen, Meclofenamic acid, Ibuprofen and Fnoprofen) which are commercially available as oral preparations or injections were tested. The cytotoxicity of 5 chemicals was evaluated by using MTT[tetrazolium salt 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide] assay. NRU (neutral red uptake) assay and Alamar Blue assay after fibroblast cells had been exposed to the chemicals for 24 hours or 489 hours. The $IC_{50}$ values of the chemicals showed the comparative strength of cytotoxicity as following order of Meclofenamic acid>Diclofenac>Fenoprofen>Ibuprofen>Naproxen. The values of $IC_{50}$ determined by Alamar Blue assay were lower than those of MTT and NRU assay. These data suggest Alamar Blue assay can be useful method for assessing in vitro skin irritation potential of anti-inflammatory drugs.

• 한국어병학회지
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• 제14권1호
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• pp.3-10
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• 2001

식물성 생약재의 항균력과 활성산소(superoxide anion, $O_2^-$)의 생산에 미치는 in vitro 효과를 조사하여 금후 in vivo 실험에 적용할 때, 어류 체내 자연 저항성 증진을 꾀할 수 있는 식물성 생약재를 발굴하기 위한 기초자료를 얻고자 하였다. 식물성 생약재는 고삼, 귀전우, 구절초, 박하, 방풍, 연교, 약쑥, 지유, 삼백초, 삼지구엽초, 상백피, 소회향, 편축, 팔각 등 14종류로 이들을 열수추출하여 실험에 사용하였다. 항균력 시험에는 어병세균 Listonella anguillarum(Vibrio anguillarum), Vibrio sp., Vibrio alginolyticus, Edwardsiella tarda, Streptococcus sp., Lactcoccus garvieae를 대상으로 disk법을 사용하였으며, 활성산소의 생산능은 넙치 신장 대식세포를 이용한 nitroblue tetrazolium(NBT) 반응으로써 조사하였다. 그 결과,약쑥이 다른 생약재에 비하여 시험에 사용된 어병 세균에 대한 항균활성이 월등히 뛰어났으며, 또한 넙치 대식세포의 활성산소 생산능을 크게 자극하는 효과가 있음을 확인할 수 있었다.

• 치과방사선
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• 제27권1호
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• pp.107-122
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• 1997

Radiation sensitivity data was generated for two human cancer cell lines(KB, RPMI 2650) and human primary gingival fibroblast was tested three times using a viable cell number counting with a hemocytometer, MTT(3-[4,5-Dimethylthiazol2-yl]-2,5-diphenyl tetrazolium bromide) assay, and LDH(Lactate dehydrogenase) assay. Single irradiation of 2, 4, 6, 10, 15, 20Gy were applied to the tumor cell lines and the primary cultured gingival fibroblast The two fractions of 4Gy and 10Gy were seperated with a 4 hour time interval. The irradiation was done with 241.5cGy/min dose rate using /sup 137/Cs MK cell irradiator at room temperature. The obtained results were as followed : 1. There was significantly different viable cell numbers as the amount of radiation dose on the tested cells were cell number counted with a hemocytometer. In fractions, there were more viable cells remaining. 2. Phase-contrast microscopically, radiation-induced morphologic changes were pronounced on the tumor cells, however, almost no differences on the gingival fibroblast. 3. There was significantly different absorbance at 2Gy on RPMI 2600, 4Gy on KB and GF in MTT assay. In fractions, the absorbance was significantly higher on KB. 4. The level of extracellular LDH activity in the experimental group was significantly higher in the 2-4Gy than the control group. 5. The total level of extracellular and intracellular LDH activity was decreased as increased amounts of radiation dose was applied.

• Toxicological Research
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• 제11권2호
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• pp.241-246
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• 1995

This study was carried out to develop the antitoxic compound about cytotoxicity of cadmium on cultured rat neuroglial cells. These cells divided into 3 groups; control group (medium only) or $MTT_{50}$ group (neuroglial cell, $61{\mu}M$ cadmium) and experimental group ($61{\mu}M$ caffeic acid). Neutral red (NR) and tetrazolium MTT of the colorimetric assay were performed to evaluate the cytotoxicity of cell organelles. The light microscopic study was carried out to morphological changes of cultured rat neuroglical cells. The results indicated that caffeic acid showed detoxification effect on cytotoxicity of cadmium in $61{\mu}M$ concentration. According to the spectroscopic study of 1:1 complex of cadmium and caffeic acid, it showed that this formation of complex eliminated cadmium from cultured rat neurogllal cells.

• 한국동물위생학회지
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• 제18권2호
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• pp.163-176
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• 1995

Triphenyl Tetrazolium Chloride(TCC) reduction test is simple and sensitive to some residual antibiotics (especially to penicillin) in milk, but comparatively insensible to sulfo-namides. The volumn of sample is also large. Thus this study was undertaken to increase the detectable level of sulfonamides in raw milk. In this study, we used small transparent plastic hole and pulp disc instead of 10m1 test tube and made test medium in which was added 0.08%TTC, 0.3% agar, 10% skim milk, approximately $10^6$ CFU/ml streptococcus thermophilus and 5ppm Trimethoprim to enhance the sensitivity for sulfonamides The results of TCC reduction test by disc plate method were summarized as follows : 1. sensitivity to residual sulfonamides were much higher than official TCC reduction test. Detectable limites of sulfamethazine, sulfamerazine, sulfathiazole, sulfachloropy-ridazine, sulfadimethoxine, sulfamononethoxine, sulfadiazine and sulfaquinoxaline were 0.1-0.5ppm levels. 2. Detectable limites to some antibiotics were simillar or good than that of official method as 0.005-0.1ppm to three ${\beta}$ -lactams, 0.25-0.5ppm to one macrolide, 2-10ppm to three aminoglycosides, 0.2-0.5ppm to three tetracycline, 0.1-0.5ppm to chloramphenicol. 3. Only 0.1ml of milk was needed to test and the test medium could be stored appnoximatly 7days in the refrigerator. So test procedure was convenient than offcial method. 4. These results suggest that disc plate method is more useful to detect bacterial growth inhibition substances including sulfonamides in raw milk.

• 한국어병학회지
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• 제15권2호
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• pp.65-75
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• 2002

This study was performed to know the effects of stress induced by the daily fluctuation of water temperture from 18$^{\circ}C$ to 25$^{\circ}C$ up and down for 30 days on the defence mechanism of olive flounder. Puralichthys olivaceus. To make clear the temperature stress on the defense mechanism of the tested fish. several factors of immune response such as counting of leucocyte appearance in peripheral blood, phagocytic activity in whole blood cells, nitroblue tetrazolium(NBT) reduction, chemiluminescence(CL) response, and lysozyme activity were investigated at 28 days after giving the change of water temperature. The fish was controlled under the none feeding condition during experimental period. Mortality of the tested fish was rapidly increased up to 22% within the first one week of the experimental period without any additional stress factors. The number of neutrophil of peripheral blood in the tested group was significantly higher than the control group at the 2nd week. but the number of lymphocyte was significantly lower than the control group at the 1st and 3rd day of the experimental period. respectively. In the NBT reduction test, the activity of macrophage in the control group fish was the highest on the 7th day while that in the tested group was on the 3rd day. Also. the phagocytosis of tested group against formalin killed cells was retarded compared with the control. CL response of the tested group was significantly lower from 2nd to 5lh day of the experimental period than the control. 'The lysozyme activity of tested group was remained higher during the experimental period than the control. Even though the tested fish showed different results in some non-specific factors of immune respceses between tested and control group fish, olive flounder seems highly adaptable in repealed water temperature change in condition after one week under the given temperature fluctuation range.

• Biomolecules & Therapeutics
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• 제20권6호
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• pp.562-568
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• 2012

Cytochrome P450 BM3 (CYP102A1) from Bacillus megaterium is a self-sufficient monooxygenase that consists of a heme domain and FAD/FMN-containing reductase domain (BMR). In this report, the reduction of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and 5-cyano-2,3-ditolyl tetrazolium chloride (CTC) by BMR was evaluated as a method for monitoring BMR activity. The electron transfer proceeds from NADPH to BMR and then to BMR substrates, MTT and CTC. MTT and CTC are monotetrazolium salts that form formazans upon reduction. The reduction of MTT and CTC followed classical Michaelis-Menten kinetics ($k_{cat}=4120\;min^{-1}$, $K_m=77{\mu}M$ for MTT and $k_{cat}=6580\;min^{-1}$, $K_m=51{\mu}M$ for CTC). Our continuous assay using MTT and CTC allows the simple, rapid measurement of BMR activity. The BMR was able to metabolize mitomycin C and doxorubicin, which are anticancer drug substrates for CPR, producing the same metabolites as those produced by CPR. Moreover, the BMR was able to interact with CYP1A2 and transfer electrons to promote the oxidation reactions of substrates by CYP1A2 and CYP2E1 in humans. The results of this study suggest the possibility of the utilization of BMR as a surrogate for mammalian CPR.