• Proceedings of the PSK Conference
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• 2003.10b
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• pp.142.1-142.1
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• 2003

Sensitive and safe method for visualization of DNA in agarose gels using visible dye is described. To improve the sensitivity, we studied a counterion-dye staining method using methyl orange as a counterion-dye which contributes to reduce excessive background staining by indoine blue. Dye concentrations, PH of staining solution, mixing molar ratio of two dyes, and staining times were optimized for the counterion-dye staining. By the staining with a mixed solution of 0.005% indoine blue and 0.00165% methyl orange in 10% ethanol 0.2M sodium acetate, 8 ng of the 3 kb DNA in an agarose gel was detected within 1hr. (omitted)

• Korean Journal of Clinical Laboratory Science
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• v.48 no.4
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• pp.371-377
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• 2016

Decalcification is routinely performed to obtain a pathological diagnosis using bone marrow biopsy. During the decalcification process using a conventional acidic solution, such as HCl, the antigenicity of tissue is damaged. Especially DNA and RNA in the bone marrow are impaired. Hence, there is the need for a standardized decalcification protocol that preserves the antigenicity of tissue. To this end, we compared the effects of two commonly used decalcifiers: Commercial decalcifier (Calcl-Clear Rapid, HCl) and the EDTA (12.5%, pH 7.0). Bone marrow biopsies sampled from 71 patients were decalcified in accordance with the protocols of respective groups-HCI versus EDTA. The differences of decalcification protocols were analyzed with respect to Hematoxylin & Eosin staining, Gomori'sreticulum staining, and immunohistochemical staining and molecular analysis. Immunohistochemical staining used Ki-67, CD20 and CD138 as primary antibodies and molecular analysis was conducted through the DNA concentration analysis, in situ hybridization (ISH) and immunoglobulin heavy chain (IGH) gene rearrangement. On the routine histopathology analysis, there was no difference between HCl and EDTA. Moreover, in case of immunohistochemical staining, the cytoplasmic membrane or cytoplasmic CD markers was well preserved. However, nuclear proteins, such as Ki-67, were stained with low quality. Conversely, according to the molecular analysis, the EDTA protocol preserved the DNA and RNA compared with the HCI. The differences of DNA quantity and quality were statistically significant between protocols of HCl and EDTA. We used 38 cases in HCl and 12 cases in EDTA. Consequently, the EDTA protocol maintains the antigenicity of the protein on tissue and is acceptable for examination with molecular base analysis. Decalcification of bone marrow biopsy by EDTA is highly recommended for the examination of immunohistochemical staining and molecular analysis.

• Journal of Korean Academy of Oral and Maxillofacial Radiology
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• v.21 no.2
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• pp.183-197
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• 1991

After single fraction of 2, 5, 10 Gy irradiation on submandibular gland of 40 male rats, weighing 150gm, respectively, these animal were sacrificed two hours after 0.1㎎/g bromodeoxyuridine (Sigma) peritoneal injection in 1, 3, 7, 15 hours, 1, 3, 7 days after irradiation. And excised submandibular gland were fixed in Carnoy's and Bouin's solution for 2 hours. Paraffin sections were stained with H&E, and PAS for the observation of the change of salivary gland tissue, and with Feulgen for the study of the DNA distribution, and immunohistochemically stained with anti-bromodeoxyuridine (Sanbyo Co.) for detection of DNA synthetic cells in order to study the distribution of DNA synthetic cells of salivary gland tissue and endothelium after irradiation in 5 different sites of 6 slides on X 200 high power field. The results were as followings. 1. In PAS staining 3 days after 5Gy irradiation, decreased mucine secretion of serous cells were found, and 7 days after l0Gy irradiation, decreased mucine secretion of mucous cells were found. 2. In histopathologic features, degeneration of serous cells were found in 3 days after 2 Gy irradiation and there was little change in mucous cells and excretory duct cells. 3. In Feugen staining, 3 days after 2 Gy, 5 Gy irradiation, more high percentage of DNA synthetic cells were found in intercalated duct cells, striated duct cells and excretory duct cells than in BrdU staining. 4. In immunohistochemical features, DNA synethsis of serous cells and granular convoluted tubular cells abruptly decreased in early period after irradiation and showed no recovery in 7 days after irradiation but there was an increase in DNA synthesis of intercalated duct cells, striated duct cells and excretory duct cells, which have less S-phase cells comparatively, in 7 days after 2 Gy, 5 Gy irradiation. 5. In immunohistochemical features, the DNA synthesis of endothelial cells was continuously decreased after irradiation but showed slight increase in 7 days after 2 Gy and S Gy irradiation.

• Asian-Australasian Journal of Animal Sciences
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• v.18 no.5
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• pp.626-631
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• 2005

Nitric oxide (NO) derived from inducible nitric oxide synthase (iNOS) is involved in cell apoptosis, which contributes to luteal regression and luteolysis in some species. In large domestic animals, no direct evidence for the relationship between NO and cell apoptosis in the process of corpus luteum regression is reported. The present study was conducted to investigate the localization of iNOS on porcine corpora lutea (CL) during the oestrus cycle and its relation to cell DNA fragmentation and CL regression. According to morphology, four luteal phases throughout the estrous cycle were defined as CL1, CL2, CL3 and CL4. By isoform-specific antibody against iNOS, the immunochemial staining was determined. Luteal cell DNA fragmentation was determined by flow cytometry. The results showed that no positive staining for iNOS was in CL1 and that iNOS was produced but limited to the periphery of CL2, while in the CL3, the spreading immunochemical staining was found inside the CL. No iNOS positive staining was detected in CL4. Meanwhile, DNA fragmentation increased dramatically when CL developed from CL2 to CL3 (p<0.05). In CL4, higher proportion of luteal cells still had fragmented DNA than that of luteal cells from CL1 or CL2 (p<0.05). These results indicate that iNOS expression is closely related to luteal cell apoptosis and then to luteal regression.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.5
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• pp.2123-2128
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• 2014

A current hurdle in cancer management is the intrinsic or acquired resistance of cancer cells to chemical agents that restricts the efficacy of therapeutic strategies. Accordingly, there is an increasing desire to discover new natural compounds with selective toxicity to combat malignancies. In present study, the cytotoxic and apoptosis-inducing activities of ferutinin, a terpenoid derivative from Ferula ovina, were investigated on human breast (MCF7) and bladder (TCC) cancer cells as well as normal fibroblasts (HFF3).The toxicity and DNA damage inducing effects of ferutinin were studied by MTT and comet assays, DAPI and PI staining and DNA laddering. The $IC_{50}$ values of ferutinin were identified and compared with routine prescribed drugs, doxorubicin and vincristine, by MTT test. Alkaline comet assay and DAPI staining revealed DNA damage due to ferutinin, which was significantly (p<0.001) higher in MCF7 and TCC than HFF3 cells. Apoptosis induction was evidenced by PI staining and DNA laddering. Our results suggest that ferutinin could be considered as an effective anticancer agent for future in vivo and clinical experiments.

• KOREAN JOURNAL OF CROP SCIENCE
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• v.50 no.spc1
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• pp.228-230
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• 2005

The denature polyacrylamide gel stain silver nitrate is used for routine nucleic acid detection. More sensitive stains, such as Vistra Green, SYBR Green are available to address a broad range of DNA applications requiring lower detection limits in polyacrylamide gel formats. Gel Scanners, laser-scanning instruments, provide sensitive fluorescence detection of DNA gel stains. We established one step fluorescent impregnation enhanced sensitivity with simple, rapid and low cost. We have applied this fluorescent staining procedure for the routine analysis of DNA profiles generated by SSR amplification.

• Toxicological Research
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• v.7 no.2
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• pp.157-163
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• 1991

The effect of patulin, a mycotoxin, on the growth of Escherichia coli cell was investigated. E. coli cell elongation usually shown in SOS-response for DNA repair was induced by 20 mg of patulin per ml. After staining the E. coli chromosome with fluorescence dye(DAPI, 4', 6-diamino-2-phenyl-indole), chromosomal DNA partitioning was not affected by patulin. The observation indicateds that patulin acts as a DNA damaging agent which is effective for E. coli cell elongation introduced by the inhibition of septum formation.

• Biomolecules & Therapeutics
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• v.20 no.5
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• pp.487-491
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• 2012

The aim of this study was to investigate the inhibitory effects of rutaecarpine on DNA strand breaks and apoptosis induced by hydrogen peroxide ($H_2O_2$) in murine Hepa-1c1c7 cells. Oxidative DNA damage was estimated by nuclear condensation assessment, fluorescence-activated cell sorting analysis, and Comet assay. Rutaecarpine inhibited cell death induced by $500{\mu}M$ $H_2O_2$, as assessed by 4',6-diamidino-2-phenylindole (DAPI) staining. Treatment with rutaecarpine reduced the number of DNA strand breaks induced by $H_2O_2$, as assessed by DAPI staining and Comet assay, and increased quinone reductase, phosphatidylinositol 3-kinase, and pAkt protein levels, as assessed by western blotting.

• Asian Pacific Journal of Cancer Prevention
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• v.14 no.3
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• pp.1643-1647
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• 2013

Background: To identify the risk factors and assess the role of survivin in predicting progessivity precancerous cervical lesions. Materials and Methods: This case-control study was conducted from October 2009 until May 2010. We obtained 74 samples, classified according to the degree of cervical intraepithelial neoplasia (CIN): 19 samples for CIN 1, 18 samples for CIN 2, 18 samples for CIN 3, and 19 samples as controls. Demographic profiles and risk factors assesment, histopathologic examination, HPV DNA tests, immunocytochemistry (ICC) and immunohistochemistry (IHC) staining for survivin expression were performed on all samples. Data was analyzed with bivariate and multivariate analysis. Results: Multivariate analysis revealed significant risk factors for developing precancerous cervical lesions are age <41 years, women with ${\geq}2$ sexual partners, course of education ${\geq}13$ years, use of oral contraceptives, positive high-risk HPV DNA, and high survivin expression by ICC or IHC staining. These factors were fit to a prediction model and we obtained a scoring system to predict the progressivity of CIN lesions. Conclusions: Determination of survivin expression by immunocytochemistry staining, along with other significant risk factors, can be used in a scoring system to predict the progressivity of CIN lesions. Application of this scoring system may be beneficial in determining the action of therapy towards the patient.

• 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.