The Experts below are selected from a list of 312 Experts worldwide ranked by ideXlab platform
Marten Ferno - One of the best experts on this subject based on the ideXlab platform.
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rna quality in frozen breast cancer samples and the influence on gene expression analysis a comparison of three evaluation methods using microcapillary electrophoresis traces
BMC Molecular Biology, 2007Co-Authors: Carina Strand, Johan Enell, Ingrid Hedenfalk, Marten FernoAbstract:Assessing RNA quality is essential for gene expression analysis, as the inclusion of degraded samples may influence the interpretation of expression levels in relation to biological and/or clinical parameters. RNA quality can be analyzed by agarose gel electrophoresis, UV spectrophotometer, or microcapillary electrophoresis traces, and can furthermore be evaluated using different methods. No generally accepted recommendations exist for which technique or evaluation method is the best choice. The aim of the present study was to use microcapillary electrophoresis traces from the Bioanalyzer to compare three methods for evaluating RNA quality in 24 fresh frozen invasive breast cancer tissues: 1) Manual method = subjective evaluation of the Electropherogram, 2) Ratio Method = the ratio between the 28S and 18S peaks, and 3) RNA integrity number (RIN) method = objective evaluation of the Electropherogram. The results were also related to gene expression profiling analyses using 27K oligonucleotide microarrays, unsupervised hierarchical clustering analysis and ontological mapping.
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RNA quality in frozen breast cancer samples and the influence on gene expression analysis – a comparison of three evaluation methods using microcapillary electrophoresis traces
BMC Molecular Biology, 2007Co-Authors: Carina Strand, Johan Enell, Ingrid Hedenfalk, Marten FernoAbstract:Assessing RNA quality is essential for gene expression analysis, as the inclusion of degraded samples may influence the interpretation of expression levels in relation to biological and/or clinical parameters. RNA quality can be analyzed by agarose gel electrophoresis, UV spectrophotometer, or microcapillary electrophoresis traces, and can furthermore be evaluated using different methods. No generally accepted recommendations exist for which technique or evaluation method is the best choice. The aim of the present study was to use microcapillary electrophoresis traces from the Bioanalyzer to compare three methods for evaluating RNA quality in 24 fresh frozen invasive breast cancer tissues: 1) Manual method = subjective evaluation of the Electropherogram, 2) Ratio Method = the ratio between the 28S and 18S peaks, and 3) RNA integrity number (RIN) method = objective evaluation of the Electropherogram. The results were also related to gene expression profiling analyses using 27K oligonucleotide microarrays, unsupervised hierarchical clustering analysis and ontological mapping.
Robert H. Yolken - One of the best experts on this subject based on the ideXlab platform.
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SHORT TECHNICAL REPORTS Evaluating RNA status for RT-PCR in extracts of postmortem human brain tissue
2013Co-Authors: Christine L. Miller, Suad Diglisic, Flora Leister, Maree Webster, Robert H. YolkenAbstract:Total RNA was extracted from 105 individual postmortem human brain samples representing a range of postmortem conditions. To improve upon parameters currently used to screen for RNA quality, Electropherogram patterns generated by the Agilent Bioanalyzer 2100 were compared to the average score in random hexamer-primed reverse transcription real-time PCR for four housekeeping genes in each RNA sample. The ribosomal ratio (28S to 18S) was found to be unrelated to the housekeeping gene score (r =-0.06; P = 0.50), and there was no threshold value in the ratio that could be applied to effectively categorize the RNA degradation. Although the housekeeping gene score correlated significantly with the percentage of area in the Electropherogram corresponding to moderate to high molecular weight intact mRNA (r = 0.41; P = 0.0001), the best discriminator was determined to be the ratio of the 18S peak height to the highest peak in the tRNA to 18S rRNA baseline. Applying a lower boundary of 2.12 for the ratio allowed for the screening out of samples with the lowest housekeeping gene scores without excluding better-quality samples. This measure represents a marked improvement over the 28S to 18S ratio, which proved to be a misleading indicator of the state of the mRNA for use in random hexamer-primed reverse transcription PCR
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evaluating rna status for rt pcr in extracts of postmortem human brain tissue
BioTechniques, 2004Co-Authors: Christine L. Miller, Suad Diglisic, Flora Leister, Maree J. Webster, Robert H. YolkenAbstract:Total RNA was extracted from 105 individual postmortem human brain samples representing a range of postmortem conditions. To improve upon parameters currently used to screen for RNA quality, Electropherogram patterns generated by the Agilent Bioanalyzer 2100 were compared to the average score in random hexamer-primed reverse transcription real-time PCR for four housekeeping genes in each RNA sample. The ribosomal ratio (28S to 18S) was found to be unrelated to the housekeeping gene score (r = −0.06; P = 0.50), and there was no threshold value in the ratio that could be applied to effectively categorize the RNA degradation. Although the housekeeping gene score correlated significantly with the percentage of area in the Electropherogram corresponding to moderate to high molecular weight intact mRNA (r = 0.41; P = 0.0001), the best discriminator was determined to be the ratio of the 18S peak height to the highest peak in the tRNA to 18S rRNA baseline. Applying a lower boundary of 2.12 for the ratio allowed f...
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Evaluating RNA status for RT-PCR in extracts of postmortem human brain tissue.
BioTechniques, 2004Co-Authors: Christine L. Miller, Suad Diglisic, Flora Leister, Maree J. Webster, Robert H. YolkenAbstract:Total RNA was extracted from 105 individual postmortem human brain samples representing a range of postmortem conditions. To improve upon parameters currently used to screen for RNA quality, Electropherogram patterns generated by the Agilent Bioanalyzer 2100 were compared to the average score in random hexamer-primed reverse transcription real-time PCR for four housekeeping genes in each RNA sample. The ribosomal ratio (28S to 18S) was found to be unrelated to the housekeeping gene score (r = -0.06; P = 0.50), and there was no threshold value in the ratio that could be applied to effectively categorize the RNA degradation. Although the housekeeping gene score correlated significantly with the percentage of area in the Electropherogram corresponding to moderate to high molecular weight intact mRNA (r = 0.41; P = 0.0001), the best discriminator was determined to be the ratio of the 18S peak height to the highest peak in the tRNA to 18S rRNA baseline. Applying a lower boundary of 2.12 for the ratio allowed for the screening out of samples with the lowest housekeeping gene scores without excluding better-quality samples. This measure represents a marked improvement over the 28S to 18S ratio, which proved to be a misleading indicator of the state of the mRNA for use in random hexamer-primed reverse transcription PCR.
Carina Strand - One of the best experts on this subject based on the ideXlab platform.
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rna quality in frozen breast cancer samples and the influence on gene expression analysis a comparison of three evaluation methods using microcapillary electrophoresis traces
BMC Molecular Biology, 2007Co-Authors: Carina Strand, Johan Enell, Ingrid Hedenfalk, Marten FernoAbstract:Assessing RNA quality is essential for gene expression analysis, as the inclusion of degraded samples may influence the interpretation of expression levels in relation to biological and/or clinical parameters. RNA quality can be analyzed by agarose gel electrophoresis, UV spectrophotometer, or microcapillary electrophoresis traces, and can furthermore be evaluated using different methods. No generally accepted recommendations exist for which technique or evaluation method is the best choice. The aim of the present study was to use microcapillary electrophoresis traces from the Bioanalyzer to compare three methods for evaluating RNA quality in 24 fresh frozen invasive breast cancer tissues: 1) Manual method = subjective evaluation of the Electropherogram, 2) Ratio Method = the ratio between the 28S and 18S peaks, and 3) RNA integrity number (RIN) method = objective evaluation of the Electropherogram. The results were also related to gene expression profiling analyses using 27K oligonucleotide microarrays, unsupervised hierarchical clustering analysis and ontological mapping.
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RNA quality in frozen breast cancer samples and the influence on gene expression analysis – a comparison of three evaluation methods using microcapillary electrophoresis traces
BMC Molecular Biology, 2007Co-Authors: Carina Strand, Johan Enell, Ingrid Hedenfalk, Marten FernoAbstract:Assessing RNA quality is essential for gene expression analysis, as the inclusion of degraded samples may influence the interpretation of expression levels in relation to biological and/or clinical parameters. RNA quality can be analyzed by agarose gel electrophoresis, UV spectrophotometer, or microcapillary electrophoresis traces, and can furthermore be evaluated using different methods. No generally accepted recommendations exist for which technique or evaluation method is the best choice. The aim of the present study was to use microcapillary electrophoresis traces from the Bioanalyzer to compare three methods for evaluating RNA quality in 24 fresh frozen invasive breast cancer tissues: 1) Manual method = subjective evaluation of the Electropherogram, 2) Ratio Method = the ratio between the 28S and 18S peaks, and 3) RNA integrity number (RIN) method = objective evaluation of the Electropherogram. The results were also related to gene expression profiling analyses using 27K oligonucleotide microarrays, unsupervised hierarchical clustering analysis and ontological mapping.
Andrew Proctor - One of the best experts on this subject based on the ideXlab platform.
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application of sequential paired covariance to liquid chromatography mass spectrometry data enhancements in both the signal to noise ratio and the resolution of analyte peaks in the chromatogram
Journal of Chromatography A, 1997Co-Authors: David C Muddiman, Alan L Rockwood, Andrew Proctor, Baoming M Huang, Gordon A Anderson, Steven A Hofstadler, Mary S Weirlipton, Richard D SmithAbstract:Abstract The algorithm of sequential paired covariance (SPC) has been previously reported to dramatically enhance the signal-to-noise (S/N) ratio for on-line separations combined with mass spectrometry. That initial study focused on a limited number of data sets derived from the combination of capillary electrophoresis (CE) with time-of-flight mass spectrometry using an electrospray interface. Results from the initial study clearly demonstrated that a significant enhancement (almost two orders of magnitude) in the S/N ratio of the eluting peaks in the Electropherogram could be obtained, facilitating identification of the analytes. In this report, the algorithm has been applied to liquid chromatography-mass spectrometry data obtained on a triple quadrupole instrument and we have evaluated the general applicability of the SPC approach to several types of microcolumn separations with mass spectrometric detection, including CE coupled with Fourier transform ion cyclotron resonance mass spectrometry. In all the cases we tested, we found the algorithm enhanced the S/N ratios of the resulting chromatograms or Electropherograms to a similar extent. This report further demonstrates the SPC approach to enhance the resolution as well as the S/N ratio of the eluting peaks of a complex peptide mixture. While many variations of the algorithm are possible, we have also found higher order covariance (e.g., 3rd order) is useful for eliminating coincidental noise in sequential mass spectra, giving the potential to extract broad, low intensity analyte peaks. We also demonstrate the sequential covariance approach for enhancing the S/N ratio of mass spectra.
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application of sequential paired covariance to capillary electrophoresis electrospray ionization time of flight mass spectrometry unraveling the signal from the noise in the Electropherogram
Analytical Chemistry, 1995Co-Authors: David C Muddiman, Alan L Rockwood, Quanyin Gao, Joanne C Severs, Harold R Udseth, Richard D Smith, Andrew ProctorAbstract:The combination of capillary electrophoresis (CE) with electrospray ionization (ESI) time-of-flight mass spectrometry (TOF-MS) has recently been demonstrated. When CE is combined with TOF-MS using an electrospray interface, the method provides fast, high-resolution separations with rapid mass analysis and the potential for very high sensitivity. In analyzing the data, one first reconstructs an Electropherogram from the data set and then identifies peaks in the mass spectra. The mass spectra corresponding to the peaks in the Electropherogram are then inspected to enable solute identification. However, background noise often prevents detection of peaks in the reconstructed Electropherogram, thus interfering with the analysis. In this work we demonstrate alternative Electropherogram reconstruction techniques that enhance its signal-to-noise ratio, allowing more immediate identification of the number of components in a mixture and their corresponding retention times. The techniques described here should also be adaptable for on-line analysis of CE-ESI-TOF data and the combination with other separation techniques coupled to mass spectrometry (e.g., LC/MS), as well as any multichannel detection scheme. 21 refs., 4 figs., 1 tab.
Christine L. Miller - One of the best experts on this subject based on the ideXlab platform.
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SHORT TECHNICAL REPORTS Evaluating RNA status for RT-PCR in extracts of postmortem human brain tissue
2013Co-Authors: Christine L. Miller, Suad Diglisic, Flora Leister, Maree Webster, Robert H. YolkenAbstract:Total RNA was extracted from 105 individual postmortem human brain samples representing a range of postmortem conditions. To improve upon parameters currently used to screen for RNA quality, Electropherogram patterns generated by the Agilent Bioanalyzer 2100 were compared to the average score in random hexamer-primed reverse transcription real-time PCR for four housekeeping genes in each RNA sample. The ribosomal ratio (28S to 18S) was found to be unrelated to the housekeeping gene score (r =-0.06; P = 0.50), and there was no threshold value in the ratio that could be applied to effectively categorize the RNA degradation. Although the housekeeping gene score correlated significantly with the percentage of area in the Electropherogram corresponding to moderate to high molecular weight intact mRNA (r = 0.41; P = 0.0001), the best discriminator was determined to be the ratio of the 18S peak height to the highest peak in the tRNA to 18S rRNA baseline. Applying a lower boundary of 2.12 for the ratio allowed for the screening out of samples with the lowest housekeeping gene scores without excluding better-quality samples. This measure represents a marked improvement over the 28S to 18S ratio, which proved to be a misleading indicator of the state of the mRNA for use in random hexamer-primed reverse transcription PCR
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evaluating rna status for rt pcr in extracts of postmortem human brain tissue
BioTechniques, 2004Co-Authors: Christine L. Miller, Suad Diglisic, Flora Leister, Maree J. Webster, Robert H. YolkenAbstract:Total RNA was extracted from 105 individual postmortem human brain samples representing a range of postmortem conditions. To improve upon parameters currently used to screen for RNA quality, Electropherogram patterns generated by the Agilent Bioanalyzer 2100 were compared to the average score in random hexamer-primed reverse transcription real-time PCR for four housekeeping genes in each RNA sample. The ribosomal ratio (28S to 18S) was found to be unrelated to the housekeeping gene score (r = −0.06; P = 0.50), and there was no threshold value in the ratio that could be applied to effectively categorize the RNA degradation. Although the housekeeping gene score correlated significantly with the percentage of area in the Electropherogram corresponding to moderate to high molecular weight intact mRNA (r = 0.41; P = 0.0001), the best discriminator was determined to be the ratio of the 18S peak height to the highest peak in the tRNA to 18S rRNA baseline. Applying a lower boundary of 2.12 for the ratio allowed f...
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Evaluating RNA status for RT-PCR in extracts of postmortem human brain tissue.
BioTechniques, 2004Co-Authors: Christine L. Miller, Suad Diglisic, Flora Leister, Maree J. Webster, Robert H. YolkenAbstract:Total RNA was extracted from 105 individual postmortem human brain samples representing a range of postmortem conditions. To improve upon parameters currently used to screen for RNA quality, Electropherogram patterns generated by the Agilent Bioanalyzer 2100 were compared to the average score in random hexamer-primed reverse transcription real-time PCR for four housekeeping genes in each RNA sample. The ribosomal ratio (28S to 18S) was found to be unrelated to the housekeeping gene score (r = -0.06; P = 0.50), and there was no threshold value in the ratio that could be applied to effectively categorize the RNA degradation. Although the housekeeping gene score correlated significantly with the percentage of area in the Electropherogram corresponding to moderate to high molecular weight intact mRNA (r = 0.41; P = 0.0001), the best discriminator was determined to be the ratio of the 18S peak height to the highest peak in the tRNA to 18S rRNA baseline. Applying a lower boundary of 2.12 for the ratio allowed for the screening out of samples with the lowest housekeeping gene scores without excluding better-quality samples. This measure represents a marked improvement over the 28S to 18S ratio, which proved to be a misleading indicator of the state of the mRNA for use in random hexamer-primed reverse transcription PCR.