Cost-effective and rapid lysis ofSaccharomyces cerevisiaecells for quantitative western blot analysis of proteins, including phosphorylated eIF2α

Yeast ◽  
2017 ◽  
Vol 34 (9) ◽  
pp. 371-382 ◽  
Author(s):  
Su Jung Lee ◽  
Rashmi Ramesh ◽  
Valerie de Boor ◽  
Jan M. Gebler ◽  
Richard C. Silva ◽  
...  
Keyword(s):  
Western Blot ◽  
Blot Analysis ◽  
Western Blot Analysis ◽  
Cost Effective ◽  
Quantitative Western Blot ◽  
Phosphorylated Eif2α

Quantitative Western blot analysis and spot immunodetection using time-resolved fluorometry

1992 ◽  
Vol 147 (2) ◽  
pp. 251-259 ◽  
Author(s):  
Eleftherios P. Diamandis ◽  
Theodore K. Christopoulos ◽  
Courtney C. Bean
Keyword(s):  
Western Blot ◽  
Blot Analysis ◽  
Western Blot Analysis ◽  
Time Resolved ◽  
Quantitative Western Blot

scholarly journals A systematic approach to quantitative Western blot analysis

2020 ◽  
Vol 593 ◽  
pp. 113608 ◽  
Author(s):  
Lakshmi Pillai-Kastoori ◽  
Amy R. Schutz-Geschwender ◽  
Jeff A. Harford
Keyword(s):  
Western Blot ◽  
Blot Analysis ◽  
Western Blot Analysis ◽  
Systematic Approach ◽  
Quantitative Western Blot

Quantitative Western blot analysis of plasma ADAMTS13 antigen in patients with Upshaw-Schulman syndrome

2007 ◽  
Vol 120 (3) ◽  
pp. 381-386 ◽  
Author(s):  
Hiromichi Ishizashi ◽  
Hideo Yagi ◽  
Masanori Matsumoto ◽  
Kenji Soejima ◽  
Tomohiro Nakagaki ◽  
...  
Keyword(s):  
Western Blot ◽  
Blot Analysis ◽  
Western Blot Analysis ◽  
Quantitative Western Blot

Detection of the anti-Hu antibody in the serum of patients with small cell lung cancer?A quantitative western blot analysis

1990 ◽  
Vol 27 (5) ◽  
pp. 544-552 ◽  
Author(s):  
Josep Dalmau ◽  
Henry M. Furneaux ◽  
Richard J. Gralla ◽  
Mark G. Kris ◽  
Jerome B. Posner
Keyword(s):  
Lung Cancer ◽  
Western Blot ◽  
Small Cell Lung Cancer ◽  
Cell Lung Cancer ◽  
Blot Analysis ◽  
Western Blot Analysis ◽  
Small Cell ◽  
Small Cell Lung ◽  
Quantitative Western Blot

scholarly journals Microfluidic Cell Culture and Its Application in High-Throughput Drug Screening

2010 ◽  
Vol 16 (1) ◽  
pp. 101-111 ◽  
Author(s):  
Xiaojing Su ◽  
Edmond W. K. Young ◽  
Heather A. S. Underkofler ◽  
Timothy J. Kamp ◽  
Craig T. January ◽  
...  
Keyword(s):  
Western Blot ◽  
Drug Screening ◽  
Membrane Trafficking ◽  
Blot Analysis ◽  
Western Blot Analysis ◽  
Cost Effective ◽  
Microfluidic Channels ◽  
Membrane Expression ◽  
Screening Assay ◽  
Drug Induced

Evaluation of drug cardiotoxicity is essential to the safe development of novel pharmaceuticals. Assessing a compound’s risk for prolongation of the surface electrocardiographic QT interval and hence risk for life-threatening arrhythmias is mandated before approval of nearly all new pharmaceuticals. QT prolongation has most commonly been associated with loss of current through hERG ( human ether-a-go-go related gene) potassium ion channels due to direct block of the ion channel by drugs or occasionally by inhibition of the plasma membrane expression of the channel protein. To develop an efficient, reliable, and cost-effective hERG screening assay for detecting drug-mediated disruption of hERG membrane trafficking, the authors demonstrate the use of microfluidic-based systems to improve throughput and lower cost of current methods. They validate their microfluidics array platform in polystyrene (PS), cyclo-olefin polymer (COP), and polydimethylsiloxane (PDMS) microchannels for drug-induced disruption of hERG trafficking by culturing stably transfected HEK cells that overexpressed hERG (WT-hERG) and studying their morphology, proliferation rates, hERG protein expression, and response to drug treatment. Results show that WT-hERG cells readily proliferate in PS, COP, and PDMS microfluidic channels. The authors demonstrated that conventional Western blot analysis was possible using cell lysate extracted from a single microchannel. The Western blot analysis also provided important evidence that WT-hERG cells cultured in microchannels maintained regular (well plate-based) expression of hERG. The authors further show that experimental procedures can be streamlined by using direct in-channel immunofluorescence staining in conjunction with detection using an infrared scanner. Finally, treatment of WT-hERG cells with 5 different drugs suggests that PS (and COP) microchannels were more suitable than PDMS microchannels for drug screening applications, particularly for tests involving hydrophobic drug molecules.

Radiosensitivity of glioma to Gamma Knife treatment enhanced in vitro and in vivo by RNA interfering Ku70 that is mediated by a recombinant adenovirus

2010 ◽  
Vol 113 (Special_Supplement) ◽  
pp. 228-235 ◽  
Author(s):  
Qiang Jia ◽  
Yanhe Li ◽  
Desheng Xu ◽  
Zhenjiang Li ◽  
Zhiyuan Zhang ◽  
...  
Keyword(s):  
Western Blot ◽  
Gamma Knife ◽  
Blot Analysis ◽  
Western Blot Analysis ◽  
Glioma Cells ◽  
C6 Glioma ◽  
C6 Glioma Cells ◽  
C6 Cells

Object The authors sought to evaluate modification of the radiation response of C6 glioma cells in vitro and in vivo by inhibiting the expression of Ku70. To do so they investigated the effect of gene transfer involving a recombinant replication-defective adenovirus containing Ku70 short hairpin RNA (Ad-Ku70shRNA) combined with Gamma Knife treatment (GKT). Methods First, Ad-Ku70shRNA was transfected into C6 glioma cells and the expression of Ku70 was measured using Western blot analysis. In vitro, phenotypical changes in C6 cells, including proliferation, cell cycle modification, invasion ability, and apoptosis were evaluated using the MTT (3′(4,5-dimethylthiazol-2-yl)2,5-diphenyltetrazolium bromide) assay, Western blot analysis, and cell flow cytometry. In vivo, parental C6 cells transfected with Ad-Ku70shRNA were implanted stereotactically into the right caudate nucleus in Sprague-Dawley rats. After GKS, apoptosis was analyzed using the TUNEL (terminal deoxynucleotidyl transferase–mediated deoxyuridine triphosphate nick-end labeling) method. The inhibitory effects on growth and invasion that were induced by expression of proliferating cell nuclear antigen and matrix metalloproteinase–9 were determined using immunohistochemical analyses. Results The expression of Ku70 was clearly inhibited in C6 cells after transfection with Ad-Ku70shRNA. In vitro following transfection, the C6 cells showed improved responses to GKT, including suppression of proliferation and invasion as well as an increased apoptosis index. In vivo following transfection of Ad-Ku70shRNA, the therapeutic efficacy of GKT in rats with C6 gliomas was greatly enhanced and survival times in these animals were prolonged. Conclusions Our data support the potential for downregulation of Ku70 expression in enhancing the radiosensitivity of gliomas. The findings of our study indicate that targeted gene therapy–mediated inactivation of Ku70 may represent a promising strategy in improving the radioresponsiveness of gliomas to GKT.

Synthesis and Characterization of Quinoline-3-Carboxamide Derivatives as Inhibitors of the ATM Kinase

2020 ◽  
Vol 20 (23) ◽  
pp. 2070-2079
Author(s):  
Srimadhavi Ravi ◽  
Sugata Barui ◽  
Sivapriya Kirubakaran ◽  
Parul Duhan ◽  
Kaushik Bhowmik
Keyword(s):  
Western Blot ◽  
Cancer Cells ◽  
Cell Line ◽  
Cell Lines ◽  
Cancer Cell ◽  
Blot Analysis ◽  
Western Blot Analysis ◽  
Cancer Cell Lines ◽  
Atm Kinase ◽  
Cytotoxicity Studies

Background: The importance of inhibiting the kinases of the DDR pathway for radiosensitizing cancer cells is well established. Cancer cells exploit these kinases for their survival, which leads to the development of resistance towards DNA damaging therapeutics. Objective: In this article, the focus is on targeting the key mediator of the DDR pathway, the ATM kinase. A new set of quinoline-3-carboxamides, as potential inhibitors of ATM, is reported. Methods: Quinoline-3-carboxamide derivatives were synthesized and cytotoxicity assay was performed to analyze the effect of molecules on different cancer cell lines like HCT116, MDA-MB-468, and MDA-MB-231. Results: Three of the synthesized compounds showed promising cytotoxicity towards a selected set of cancer cell lines. Western Blot analysis was also performed by pre-treating the cells with quercetin, a known ATM upregulator, by causing DNA double-strand breaks. SAR studies suggested the importance of the electron-donating nature of the R group for the molecule to be toxic. Finally, Western-Blot analysis confirmed the down-regulation of ATM in the cells. Additionally, the PTEN negative cell line, MDA-MB-468, was more sensitive towards the compounds in comparison with the PTEN positive cell line, MDA-MB-231. Cytotoxicity studies against 293T cells showed that the compounds were at least three times less toxic when compared with HCT116. Conclusion: In conclusion, these experiments will lay the groundwork for the evolution of potent and selective ATM inhibitors for the radio- and chemo-sensitization of cancer cells.

The Antitumor Effects of Britanin on Hepatocellular Carcinoma Cells and its Real-Time Evaluation by In Vivo Bioluminescence Imaging

2020 ◽  
Vol 20 (9) ◽  
pp. 1147-1156
Author(s):  
Hanrui Li ◽  
GeTao Du ◽  
Lu Yang ◽  
Liaojun Pang ◽  
Yonghua Zhan
Keyword(s):  
Hepatocellular Carcinoma ◽  
Western Blot ◽  
Blot Analysis ◽  
Tumor Size ◽  
Hepg2 Cells ◽  
Western Blot Analysis ◽  
Bioluminescence Imaging ◽  
Antitumor Effects ◽  
In Vivo Bioluminescence Imaging

Background: Hepatocellular carcinoma is cancer with many new cases and the highest mortality rate. Chemotherapy is the most commonly used method for the clinical treatment of hepatocellular carcinoma. Natural products have become clinically important chemotherapeutic drugs due to their great potential for pharmacological development. Many sesquiterpene lactone compounds have been proven to have antitumor effects on hepatocellular carcinoma. Objective: Britanin is a sesquiterpene lactone compound that can be considered for the treatment of hepatocellular carcinoma. The present study aimed to investigate the antitumor effect of britanin. Methods: BEL 7402 and HepG2 cells were used to study the cytotoxicity and antitumor effects of britanin. Preliminary studies on the nuclear factor kappa B pathway were conducted by western blot analysis. A BEL 7402-luc subcutaneous tumor model was established for the in vivo antitumor studies of britanin. In vivo bioluminescence imaging was conducted to monitor changes in tumor size. Results: The results of the cytotoxicity analysis showed that the IC50 values for britanin in BEL 7402 and HepG2 cells were 2.702μM and 6.006μM, respectively. The results of the colony formation demonstrated that the number of cells in a colony was reduced significantly after britanin treatment. And the results of transwell migration assays showed that the migration ability of tumor cells was significantly weakened after treatment with britanin. Tumor size measurements and staining results showed that tumor size was inhibited after britanin treatment. The western blot analysis results showed the inhibition of p65 protein expression and reduced the ratio of Bcl-2/Bax after treatment. Conclusion: A series of in vitro and in vivo experiments demonstrated that britanin had good antitumor effects and provided an option for hepatocellular carcinoma treatment.

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