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Accurate Quantitation (accurate + quantitation)
Selected AbstractsVasomotion dynamics following calcium spiking depend on both cell signalling and limited constriction velocity in rat mesenteric small arteriesJOURNAL OF CELLULAR AND MOLECULAR MEDICINE, Issue 3 2008Ed VanBavel Abstract Vascular smooth muscle cell contraction depends on intracellular calcium. However, calcium-contraction coupling involves a complex array of intracellular processes. Quantitating the dynamical relation between calcium perturbations and resulting changes in tone may help identifying these processes. We hypothesized that in small arteries accurate quantitation can be achieved during rhythmic vasomotion, and questioned whether these dynamics depend on intracellular signalling or physical vasoconstriction. We studied calcium-constriction dynamics in cannulated and pressurized rat mesenteric small arteries (,300 ,m in diameter). Combined application of tetra-ethyl ammonium (TEA) and BayK8644 induced rhythmicity, consisting of regular and irregular calcium spiking and superposition of spikes. Calcium spikes induced delayed vasomotion cycles. Their dynamic relation could be fitted by a linear second-order model. The dirac impulse response of this model had an amplitude that was strongly reduced with increasing perfusion pressure between 17 and 98 mmHg, while time to peak and relaxation time were the largest at an intermediate pressure (57 mmHg: respectively 0.9 and 2.3 sec). To address to what extent these dynamics reside in intracellular signalling or vasoconstriction, we applied rhythmic increases in pressure counteracting the vasoconstriction. This revealed that calcium-activation coupling became faster when vasoconstriction was counteracted. During such compensation, a calcium impulse response remained that lasted 0.5 sec to peak activation, followed by a 1.0 sec relaxation time, attributable to signalling dynamics. In conclusion, this study demonstrates the feasibility of quantitating calcium-activation dynamics in vasomoting small arteries. These dynamics relate to both intracellular sig-nalling and actual vasoconstriction. Performing such analyses during pharmacological intervention and in genetic models provides a tool for unravelling calcium-contraction coupling in small arteries. [source] Quantification of urinary N -acetyl- S - (propionamide)cysteine using an on-line clean-up system coupled with liquid chromatography/tandem mass spectrometryJOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 4 2005Chien-Ming Li Abstract Acrylamide has been reported to be present in high-temperature processed foods and normal processed food intake could lead to significant acrylamide exposure. Acrylamide in vivo can be conjugated with glutathione in the presence of glutathione transferase. This conjugation product is further metabolized and excreted as N -acetyl- S -(propionamide)cysteine (NASPC) in the urine. NASPC could be considered a biomarker for acrylamide exposure. The objective of this study was to develop a highly specific, rapid and sensitive method to quantify urinary NASPC, serving as a biomarker for acrylamide exposure assessment. Isotope-labeled [13C3]NASPC was successfully synthesized and used as an internal standard. This urine mixture was directly analyzed using a newly developed liquid chromatographic/tandem mass spectrometric method coupled with an on-line clean-up system. The detection limit for this method was estimated as <5 µg l,1(0.4 pmol) on-column. The method was applied to measure the urinary level of NASPC in 70 apparently health subjects. The results showed that the NASPC urinary level was highly associated with smoking. Smokers had a significantly higher urinary NASPC level (135 ± 88 µg g,1 creatinine) than non-smokers (76 ± 30 µg g,1 creatinine). A highly sensitive and selective LC/MS/MS isotope dilution method was successfully established. With an on-line clean-up system, this system is capable of routine high-throughput analysis and accurate quantitation of NASPC in urine. This could be a useful tool for health surveillance for acrylamide exposure in a population for future study. Copyright © 2005 John Wiley & Sons, Ltd. [source] Analytical, Risk Assessment, and Remedial Implications Due to the Co-Presence of Polychlorinated Biphenyls and Terphenyls at Inactive Hazardous Waste SitesREMEDIATION, Issue 1 2000James J. Pagano Investigations conducted at three inactive hazardous waste sites in New York State have confirmed the co-presence of polychlorinated hiphenyls (PCBs) and polychlorinated terphenyls (PCTs) in soils, sediments, and biota. The PCTs at all three sites were positively identified as Aroclor 5432, with the most probable source being the hydraulic fluid Pydraul 312A utilized for high-temperature applications. The identification of the lower-chlorinated PCT formulations in environmental samples is problematical, since PCT Aroclors 5432 and 5442 are not chromatographically distinct from the higher-chlorinated (PCB) Aroclors 1254, 1260, 1262, and 1268 using conventional gas chromatography,electron capture detection. Results from this study indicate that U.S. Environmental Protection Agency (USEPA) approved PCB methods routinely utilized by most commercial laboratories based on Florisil adsorption column chromatography cleanup are inadequate to produce valid chromatographic separation and quantitative results with soils, sediment, and biota samples containing both PCBs and PCTs. The presence of co-eluting PCBs and PCTs precludes accurate quantitation due to significant differences in PCB/PCT electron capture detector response factors, and the potential for misidentification of PCT Aroclors as higher chlorinated PCB Aroclors. A method based on alumina column adsorption chromatography was used, allowing for the accurate identification and quantitation of PCB and PCT Aroclors. The results of this study suggest that the utilization of alumina adsorption column separation may have applicability and regulatory significance to other industrially contaminated sites which historically used Pydraul 312A. Inferences. [source] Relative concentrations of hK2/PSA mRNA in benign and malignant prostatic tissueTHE PROSTATE, Issue 4 2005Susanna Lintula Abstract BACKGROUND Prostate-specific antigen (PSA/KLK3) and human kallikrein 2 (hK2/KLK2) belong to the human kallikrein gene family. These two highly homologous genes are specifically expressed in the prostate under androgen control. Expression of these is regulated by similar mechanisms but changes in their relative expression have been observed in prostate cancer. METHODS We determined the relative levels of PSA and hK2 mRNA in benign and malignant prostate tissue using a quantitative reverse transcription-polymerase chain reaction (RT-PCR) method. The mRNA of PSA and hK2 are reverse transcribed and amplified in one reaction with the same primers. RESULTS The variation in the ratio of hK2/PSA mRNA was remarkably small, the difference between the highest and lowest values being three-fold. The ratio was significantly higher in WHO grade 2 compared to normal or benign prostatic hyperplasia tissue (P,=,0.032 and P,=,0.035, respectively) and in grade 3 compared to normal or benign prostatic hyperplasia tissue (P,=,0.006 in both). CONCLUSIONS The new quantitative RT-PCR technique facilitates very accurate quantitation of the relative mRNA levels of homologous genes. Using this method we have shown that the ratio of hK2/PSA mRNA is higher in cancerous than in benign prostatic tissue. © 2004 Wiley-Liss, Inc. [source] | ||||||||||