Quantitative electrophoresis in polyacrylamide gels of 2–40%

Blattler, D.P.; Garner, F. H.; Slyke, K. van; Bradley, A.R.

doi:10.1016/s0021-9673(00)92958-3

Cited by 274 publications

(88 citation statements)

References 6 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…However direct or indirect autoradiography of hydrated gels at -70°C causes a considerable loss of resolution. Gels made according to formula I of Blattler et al [6] can be dried before exposure, but if this is inconvenient, resolution can be maintained in hydrated gels by indirect autoradiography at room temperature.…”

Section: Discussionmentioning

confidence: 99%

Enhanced autoradiographic detection of ³²P and ¹²⁵I using intensifying screens and hypersensitized film

1977

View full text Add to dashboard Cite

Section: Discussionmentioning

confidence: 99%

Enhanced autoradiographic detection of ³²P and ¹²⁵I using intensifying screens and hypersensitized film

1977

View full text Add to dashboard Cite

“…Total cell proteins or immunoprecipitates were analysed by electrophoresis on 6 cm 18 % polyacrylamide gels containing 0.072 % bisacrylamide, 0.1% SDS and 3 M-urea, using the discontinuous buffer system described by Laemmli (1970). The acrylamide:bisacrylamide ratio (250:1) was determined by using the formula of Blattler et al (1972). After electrophoresis for 2-5 h at 100 V, protein gels were fixed in a solution of 20% methanol and 5% acetic acid, soaked in EN3HANCE (New England Nuclear), washed in water and dried for autoradiography.…”

Section: Methodsmentioning

confidence: 99%

Involvement of the influenza A virus PB2 protein in the regulation of viral gene expression

Mukaigawa

Hatada

Fukuda

et al. 1991

Journal of General Virology

View full text Add to dashboard Cite

To determine the function(s) of the PB2 protein of influenza A virus, six temperature-sensitive (ts) mutants of A/Udorn/72 (H3N2) virus, each carrying a ts mutation in the PB2 gene, were analysed for virus RNA and protein synthesis. One of the mutants, ICRC27, exhibited unique phenotypes and was characterized in detail. At the non-permissive temperature, 40 °C, the accumulation of mRNA for each genome segment was reduced severely, leading to delayed and reduced synthesis of viral proteins, complementary and viral RNAs (cRNAs and vRNAs). At the permissive temperature, 34 °C, the mutant virus produced severalfold greater concentrations of both mRNAs and cRNAs of PB2, PB1 and PA segments than wild-type virus. The synthesis of the three polymerase proteins and the induction of RNA polymerase activity were also greatly increased. By contrast, the expression of the haemagglutinin (HA) gene was severely suppressed. The over-production of the polymerase mRNAs was not observed during primary transcription, i.e. in the presence of cycloheximide. The ts ÷ revertants oflCRC27 did not exhibit the ts defects and also lost most of the non-ts phenotypes at 34 °C. These observations indicate that the PB2 protein participates not only in the synthesis of viral RNAs, but also in the regulation of viral gene expression, i.e. in the downregulation of the three polymerase genes and the upregulation of the HA gene during secondary transcription.

show abstract

“…Immunoblotting was performed according to Blattler et al 23 and Laemmli et al 24 Membrane fractions from PC-6, PC-6/SN2-5H2 and HEK293/ABCG2-141Lys cells (5 g of protein) were loaded onto a 7.5% SDS-PAGE gel. Following transfer to PVDF membranes, immunoblotting was performed using BXP-21 antibody (1:250; Signet Laboratories, Dedham, MA) and horseradish peroxidase-conjugated antimouse IgG (1:20,000; Kirkegaard and Perry Laboratories, Gaithersburg, MD).…”

Section: Immunoblottingmentioning

confidence: 99%

Novel camptothecin analogues that circumvent ABCG2‐associated drug resistance in human tumor cells

Yoshikawa

Ikegami

Hayasaka

et al. 2004

Intl Journal of Cancer

View full text Add to dashboard Cite

Key words: SN-38; CPT analogue; ABCG2; SAR; DNA topoisomerase I inhibitorCamptothecin (CPT) is an antitumor alkaloid that was originally isolated from Camptotheca acuminata, a tree native to southern China. 1 CPT has been demonstrated to inhibit mammalian DNA topoisomerase I (Topo I), the nuclear enzyme that changes the topologic state of duplex DNA by single-strand breakage and resealing. Stabilization of the covalent Topo I-DNA complex (so-called cleavable complex) by CPT is a critical step in its antitumor action where Topo I-mediated DNA breaks are induced via prevention of DNA relegation. 2 Clinical and preclinical studies, however, revealed reversible bone marrow depression and hemorrhagic cystitis as the major dose-limiting toxicities of CPT. Thus, efforts have been directed at finding new CPT analogues with higher antitumor activity and less toxicity.Irinotecan (7-ethyl-10-[4-(1-piperidino)-1-piperidino]-carbonyloxycamptothecin; CPT-11) has been synthesized and developed as a new water-soluble analogue of CPT with wide-spectrum antitumor activity against many types of human tumor cells. 3,4 CPT-11 is a prodrug, the antitumor activity of which is exerted by 7-ethyl-10-hydroxycamptothecin (SN-38), the active metabolite of CPT-11. 5 Despite enormous expense and efforts spent on the development of cancer chemotherapies, acquired and intrinsic drug resistance in tumors is the major obstacle to long-term sustained patient response to chemotherapy. Hitherto several mechanisms for the resistance to SN-38 and its analogues have been proposed, e.g., mutations or decreased expression of Topo I, increased expression of the UGT1A protein or single nucleotide polymorphisms (SNPs) of the UGT1A gene, increased activity of O 6 -methylguanine-DNA-methyltransferase, a DNA repair protein, decreased activity of carboxylesterase that catalyzes the biosynthesis of SN-38 from CPT-11 in the plasma and liver and overexpression of drug export pumps. 6 -11 It has been documented that several ATP-binding cassette (ABC) transporters, such as P-glycoprotein (ABCB1/MDR1/P-gp) and multidrug resistance-associated protein 1 (ABCC1/MRP1), can cause drug resistance in tumor cells by actively extruding antitumor drugs. 12,13 Recently, a novel ABC transporter, ABCG2, also known as breast cancer-resistant protein (BCRP/MXR1/ ABCP), has been discovered in drug-resistant cell lines selected for by mitoxantrone or Topo I inhibitors. 14,15 Overexpression of ABCG2 has been shown to confer resistance to doxorubicin, mitoxantrone and various CPT analogues. 14,16,17 In a recent study with plasma membrane vesicles prepared from ABCG2-overexpressing SN-38-selected human small cell lung carcinoma cells, we found that ABCG2 transported SN-38 and its glucuronide metabolite in an ATP-dependent manner. 18 Thus, it is highly likely that ABCG2 actively extrudes SN-38 from tumor cells and thereby confers drug resistance.To circumvent ABCG2-associated drug resistance, in the present study we have synthesized a total of 14 different analogues of CPT and examine...

show abstract

Quantitative electrophoresis in polyacrylamide gels of 2–40%

Cited by 274 publications

References 6 publications

Enhanced autoradiographic detection of ³²P and ¹²⁵I using intensifying screens and hypersensitized film

Enhanced autoradiographic detection of ³²P and ¹²⁵I using intensifying screens and hypersensitized film

Involvement of the influenza A virus PB2 protein in the regulation of viral gene expression

Novel camptothecin analogues that circumvent ABCG2‐associated drug resistance in human tumor cells

Contact Info

Product

Resources

About

Quantitative electrophoresis in polyacrylamide gels of 2–40%

Cited by 274 publications

References 6 publications

Enhanced autoradiographic detection of 32P and 125I using intensifying screens and hypersensitized film

Enhanced autoradiographic detection of 32P and 125I using intensifying screens and hypersensitized film

Involvement of the influenza A virus PB2 protein in the regulation of viral gene expression

Novel camptothecin analogues that circumvent ABCG2‐associated drug resistance in human tumor cells

Contact Info

Product

Resources

About

Enhanced autoradiographic detection of ³²P and ¹²⁵I using intensifying screens and hypersensitized film

Enhanced autoradiographic detection of ³²P and ¹²⁵I using intensifying screens and hypersensitized film