Loss of 5hmC identifies a new type of aberrant DNA hypermethylation in glioma.

Aberrant DNA hypermethylation is a hallmark of cancer although the underlying molecular mechanisms are still poorly understood. To study the possible role of 5-hydroxymethylcytosine (5hmC) in this process we analyzed the global and locus-specific genome-wide levels of 5hmC and 5-methylcytosine (5mC) in human primary samples from 12 non-tumoral brains and 53 gliomas. We found that the levels of 5hmC identified in non-tumoral samples were significantly reduced in gliomas.

Modification of a commercial gas chromatography isotope ratio mass spectrometer for on-line carbon isotope dilution: Evaluation of its analytical characteristics for the quantification of organic compounds.

We describe the instrumental modification of a commercial gas chromatography isotope ratio mass spectrometer (GC-IRMS) and its application for on-line carbon isotope dilution. The main modification consisted in the addition of a constant flow of enriched (13)CO2 diluted in helium after the chromatographic column through the splitter holder located inside the chromatographic oven of the instrument. In addition, and in contrast to the conventional mode of operation of GC-IRMS instruments where the signal at m/z 45 is amplified 100-fold with respect to the signal at m/z 44, the same signal amplification was used in both Faraday cups at m/z 44 and 45.

Evaluation of online carbon isotope dilution mass spectrometry for the purity assessment of synthetic peptide standards.

We present a novel method for the purity assessment of peptide standards which is applicable to any water soluble peptide. The method is based on the online (13)C isotope dilution approach in which the peptide is separated from its related impurities by liquid chromatography (LC) and the eluent is mixed post-column with a continuous flow of (13)C-enriched sodium bicarbonate. An online oxidation step using sodium persulfate in acidic media at 99°C provides quantitative oxidation to (12)CO2 and (13)CO2 respectively which is extracted to a gaseous phase with the help of a gas permeable membrane.

Liquid chromatography, chemical oxidation, and online carbon isotope dilution mass spectrometry as a universal quantification system for nonvolatile organic compounds.

A procedure for the universal detection and quantification of polar organic compounds separated by liquid chromatography (LC) based on postcolumn carbon isotope dilution mass spectrometry (IDMS) was developed. The eluent from the LC column is mixed online with a continuous flow of (13)C-enriched sodium bicarbonate, and the sodium persulfate oxidation reaction in acidic media is employed to achieve isotope equilibration. All carbon-containing compounds eluting from the column are oxidized to (12)CO(2) and (13)CO(2), respectively, and the carbon dioxide is separated from the aqueous phase using a gas-permeable membrane.

Towards compound-independent calibration for organic compounds using online isotope dilution mass spectrometry.

Isotope dilution mass spectrometry (IDMS) can be considered a primary measurement method directly traceable to the International System of Units (SI). This measurement technique is increasingly employed in routine laboratories, owing to its unequalled analytical performance, precision and ease of accreditation. Unfortunately, for the adequate application of IDMS, several isotopically labelled standards, corresponding to the compounds of interest, are required.

Gas chromatography-combustion-mass spectrometry with postcolumn isotope dilution for compound-independent quantification: its potential to assess HS-SPME procedures.

A quadrupole GC-MS instrument with an electron ionization (EI) source has been modified to enable application of postcolumn isotope dilution analysis for the standardless quantification of organic compounds injected in the gas chromatograph. Instrumental modifications included the quantitative conversion of the separated compounds into CO(2), using a postcolumn combustion furnace, and the subsequent mixing of the gas with a constant flow of (13)CO(2) diluted in helium. The online measurement of the (12)CO(2)/(13)CO(2) (44/45) ratio in the EI-MS allowed us to obtain quantitative data without resorting to compound-specific standards.

A quantitative universal detection system for organic compounds in gas chromatography with isotopically enriched (13)CO2.

Cheap and cheerful: Postcolumn carbon-isotope dilution with (13)CO(2) in combination with the coupling of gas chromatography, combustion, and EIMS (valve in position B in the picture) provides a generic quantitative approach for every organic compound without the need for specific standards. A GC-MS instrument can be upgraded by a simple low-cost modification without any loss of the structural information provided by electron ionization sources.