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At least 739 records · Page 41Linked to original sources
PubMed1980

Ultra-rapid freezing of thin biological samples.

Miniature thermocouples are used to follow the time course of cooling within test samples plunged into liquid coolants. Variations in the temperature and type of coolant (propane, Freon 22, sub-cooled liquid nitrogen), in the velocity and depth of plunge, and in the size and shape of samples were investigated. Coolants have their optimum cooling properties at their lowest liquid temperatures but show different dependencies on temperature. The optimum velocity of plunge depends on the shape of the sample. The strongest influence on cooling rate comes from the size and shape of the sample. Spherical samples about 1 mm3 show rates < 500 decrees C/s while samples 0.001 mm3 show rates > 50,000 degrees C/s for freezing in liquid propane under identical conditions. Thin samples of equivalent volume can be more rapidly frozen. Cooling rates of about 11,900 degrees C/s (0 degrees C to -100 degrees C) are recorded for thin samples (0.1 mm3 and 20 micrometers thick) sandwiched between copper strips and rapidly plunged into liquid propane (-190 degrees C). An apparatus is described for using these holders in freeze-fracture studies. Thin samples thus frozen show no evidence of ice crystal growth in either low-temperature x-ray diffraction patterns or in freeze-fracture electron micrographs.

Freeze Fracturing↗
PubMed2005

Concentration of prion protein from biological samples to increase the limits of detection by immunoassay.

An RNA-ligand-based adsorbent has been shown to concentrate prion protein (PrP) from solutions in a model system. The work presented here extends the utility of the RNA-based adsorbent to brain homogenates of cow, sheep, mule deer (Odocoileus hemionus) and elk (Cervus elaphus). Brain homogenates were diluted either in buffer, representing specimens used in post-mortem tests, or in serum, modelling specimens used in biological-fluid-based tests. The RNA adsorbent was effective in binding PrPC (cellular PrP,) and PrPres (proteinase K-resistant PrP) from the brain homogenates of all the species tested in both model systems. The three antibodies against PrP used in the experiments identified PrP in immunoblot analysis after concentrating PrP from brain homogenates with the adsorbent, indicating the general applicability of this technology for improving the detection of PrP in immunoassays. Utilization of RNA adsorbent increased the level of detection of PrPres by immunoblot over several-hundredfold. The results obtained suggest that this RNA adsorbent can be used to increase detection in current post-mortem immunoassays and for the development of a blood-based ante-mortem test.

Animals↗
PubMed2002

Simultaneous LC determination of tiazofurin, its acetyl and benzoyl esters and their active metabolite thiazole-4-carboxamide adenine dinucleotide in biological samples.

A rapid and sensitive HPLC-RP method for simultaneous determination of tiazofurin, its 5'-O acetyl and benzoyl esters and their active metabolite thiazole-4-carboxamide adenine dinucleotide was developed and validated. The method allowed determination and quantification of nanomolar quantities of these substances in cell extracts of treated cells, and was also used in kinetic studies of cellular uptake of tiazofurin and its esters from the cultivation medium. Separation of the analyzed substances from unidentified peaks from both biological materials was achieved by gradient elution, thus reducing the possibility of interference. The mobile phase consisted of a 0.1 M sodium-hydrogen phosphate, pH 5.1 and methanol. Run time was 22 min, with 5 min equilibration time.

Adenine Nucleotides↗
PubMed2001

Millimeter wave power density in aqueous biological samples.

Power density distribution inside a water sample placed between two parallel lossy dielectric plates (Polystyrene) was calculated using Fresnel equations for the frequency range of 42.25-53.57 GHz. Due to the multiple internal reflections from the sample boundaries, the distribution of the power density within the thin sample is more uniform than that within a semi-infinite medium. The power density in a sample depends on the thicknesses of the sample and the adjacent dielectric plates. For the given frequency range the sample thickness optimal for power density uniformity varies between 0.28 and 0.33 mm. The front plate has a significant effect on the magnitude of the power density within the sample but little effect on the power density distribution. The thicker the rear plate, the greater is the non uniformity of the power density distribution within the sample. Based on the calculated data, we determined the dimension of an exposure chamber providing the optimal power density distribution uniformity for mm-wave irradiation.

Air↗
PubMed2005

Supercritical fluid extraction of isoflavones from biological samples with ultra-fast high-performance liquid chromatography/mass spectrometry.

An efficient method of modifier addition for supercritical fluid extraction (SFE) of polar isoflavones was developed and yielded extraordinarily high recoveries. To find the optimal extraction conditions, a temperature and pressure optimization and modifier impact study was performed in naturally contaminated and spiked samples. Ultra-fast high-performance liquid chromatography/mass spectrometry (HPLC/MS) was used for the determination of isoflavones on an Atlantis dC18 high-speed reversed phase chromatographic column (20 x 2.1 mm, 3 microm particle size). A newly elaborated supercritical fluid extraction (SFE) procedure allowed more accurate (< 5%) and precise (< 4-7%) determination of isoflavones in biological materials. The HPLC/MS method significantly reduced analysis time with simultaneous improvement of sensitivity and detection limits. The on-column limits of detection LOD (S/N = 3) for isoflavone glycosides (daidzin, genistin, glycitin, ononin, and sissotrin) were 1.3-3.6 fmol and 0.2-1.0 fmol for aglycones (daidzein, glycitein, genistein, formononetin, and biochanin A), respectively.

Chromatography, High Pressure Liquid↗
PubMed1977

Use of octadecasilyl-silica for the extraction and purification of peptides in biological samples. Application to the identification of circulating metabolites of corticotropin-(1-24)-tetracosapeptide and somatostatin in vivo.

Peptides can be adsorbed on octadecasilyl-silica from large volumes of aqueous solution and eluted with aqueous solvent mixtures containing methanol or acetonitrile. These properties may be used for the extraction and purification of peptide fragments in plasma samples collected from rats. After intravenous injection of Synacthen [corticotropin-(1-24)-tetracosapeptide], it was shown that within 2 min the main circulating products were intact peptide and its sulphoxide. In addition, a number of fragments indicative of cleavage at the N- and C-termini were present. Most of the products formed from Synacthen have low biological activity. Somatostatin was rapidly cleaved in vivo and in vitro to a single product, which probably retains biological activity. The absence of other circulating products suggests that somatostatin is only inactivated once it leaves the circulation.

Adrenocorticotropic Hormone↗
PubMed2006

Simple linear models of scanning impedance imaging for fast reconstruction of relative conductivity of biological samples.

Scanning impedance imaging (SH) uses a noncontacting electrical probe held at a known voltage and scanned over a thin sample on a ground plane in a conductive medium to obtain images of current. The current image is related in a nonlinear way to the conductivity of the sample. This paper develops the theory behind SII showing how the measured current relates to the desired conductivity. Also included is the development of a simplified, linear model that is effective in explaining many of the experimental results. Good agreement of the linear model with step-response data over an insulator is shown. The linear model shows that the current is a blurred version of the conductivity. Simple deblurring methods can, therefore, be applied to obtain relative conductivity images from the raw current data. Raw SII data from a flower-petal and a leaf sample are shown as well as relative conductivity images deblurred using the linear model.

Algorithms↗
PubMed2006

A specific method for measurement of equine active myeloperoxidase in biological samples and in in vitro tests.

An original method called SIEFED (specific immunological extraction followed by enzymatic detection) was developed for the specific detection of the activity of equine myeloperoxidase (MPO). The method consists of the extraction of MPO from aqueous solutions by immobilized anti-MPO antibodies followed by washing (to eliminate proteins and interfering molecules) and measurement of MPO activity using a detection system containing a fluorogenic substrate, hydrogen peroxide, and nitrite as reaction enhancer. The SIEFED technique was applied to study active MPO in horse biological fluids and the effects of 2 polyphenolic molecules, curcumin and resveratrol, on MPO activity. The detection limit of the SIEFED was 0.23 mU/ml. The SIEFED exhibited good precision with intra-assay and interassay coefficients of variation below 10% and 20%, respectively, for MPO activities ranging from 0.25 to 6.4 mU/ml. The activity of MPO was generally higher than 1 mU/ml in the fluids collected from horses with inflammatory diseases. Curcumin and resveratrol exerted a dose-dependent inhibition on MPO activity and, as they were removed before the enzymatic detection of MPO, the results suggest a direct drug-nzyme interaction or an enzyme structure modification by the drug. The SIEFED is a new tool that would be useful for specific detection of active MPO in complex media and for selection of MPO activity modulators.

Animals↗
PubMed1992

A monoclonal antibody-based immunoassay for detecting tetrodotoxin in biological samples.

Spleen cells from mice hyperimmunized with a keyhole limpet hemocyanin-tetrodotoxin-formaldehyde conjugate were fused with murine P3X63Ag8.653 myeloma cells. A single hybridoma clone was identified that secretes an IgG1,k monoclonal antibody (MAb), designated T20G10, against tetrodotoxin (TTX), with an estimated affinity of 1.2 x 10(8) L/M. Competitive inhibition enzyme immunoassays (CIEIAs) for detecting TTX were developed using this MAb. A direct CIEIA using alkaline phosphatase-labeled MAb detected TTX with sensitivities at IC50 and IC20 of 6-7 ng/ml and 2-3 ng/ml, respectively. The accuracy of the direct CIEIA was comparable with the high-performance liquid chromatography (HPLC) and the mouse bioassay systems, but the direct CIEIA exhibited greater sensitivity. The direct CIEIA was also more cost effective, as it required less sample preparation, a shorter assay time, and reduced investment in equipment than either of the other assay systems.

Animals↗
PubMed1994

A survey to assess the ultrastructural preservation of fixed biological samples after air-drying from tetramethylsilane.

A scanning electron microscope (SEM) survey assessed the quality of ultrastructural preservation of a wide range of tissues from the laboratory rat and from a limited number of botanical and fly-larval samples, air-dried from tetramethylsilane (TMS) after chemical fixation and dehydration. Comparisons were made with critical point drying (CPD) and air-drying from acetone. CPD produced good results in all cases. TMS gave acceptable results with 75%, and acetone with 31%, of rat tissues. Air-drying was unacceptable for the botanical and fly-larval samples. It is concluded that air-drying from TMS is generally useful for mammalian tissues in preparation for the SEM.

Animals↗
PubMed1978

[Ambroxol, studies of biotransformation in man and determination in biological samples (author's transl)].

15 mg trans-4-[2-amino-3,5-dibromo-benzyl)-amino]-cyclohexanol-hydrochloride (ambroxol, NA 872) was administered i.v. and orally to healthy volunteers. The metabolic pattern in urine and plasma was similar for both routes of administration. Biotransformation reactions are straightforward, yielding two major products of phage I reactions identified as 6,8-dibromo-3-(trans-4-hydroxycyclohexyl)-1,2,3,4-tetrahydro-quinazoline and 3,5-dibromo-anthranilic acid. These metabolites as well as the parent compound are also converted to conjugates, predominantly glucuronides. Quantification of unlabelled ambroxol in biological fluids is achieved by radiochemical derivatisation with 14C-labelled formaldehyde in imitation of the biotransformation.

Administration, Oral↗
PubMedDate not supplied

Analysis of cholesterol oxidation products in biological samples.

Cholesterol oxidation products, or oxysterols, have gained increased attention since it was suggested that they participate in cell signaling as ligands for the nuclear receptors liver X receptor alpha and beta. In addition, oxysterols serve as important intermediates in bile acid biosynthesis and are also involved in cholesterol transport. Several studies have suggested that certain oxysterols may be used as markers for oxidative stress, and still other oxysterols may be of use in diagnosing neurological diseases. This broad spectrum of functions in health and disease has created a demand for accurate and reliable methods to measure oxysterols in complex biological matrixes. At present, the most reliable and sensitive method for oxysterol determination in biological materials is isotope-dilution gas chromatography-mass spectrometry using deuterium-labeled internal standards. With this technique, the major oxysterols in human blood plasma and atherosclerotic plaques have been carefully determined. The knowledge of oxysterol contents in other tissues and organs is still very scarce. As oxysterols are found to participate in an increasing number of cellular events, it is obvious that improved methods are needed for their analysis to understand their roles in the living cell.

Animals↗
PubMed2006

Hyperspectral NIR imaging for calibration and prediction: a comparison between image and spectrometer data for studying organic and biological samples.

A hyperspectral image in the near infrared contains thousands of position-referenced spectra. After imaging reference materials of known composition it is possible to build Partial Least Squares (PLS) regression models for predicting unknown compositions from new images or spectra. In this paper a comparison is made between spectra from a hyperspectral image and spectra from two spectrometers: a scanning grating instrument with rotating sample holders and an FT-NIR instrument utilizing a fiber-optic probe. The raw spectra and the quality of the PLS calibration models and predictions are compared. Two sample datasets consist of a set of 13 designed artificial mixtures of pure constituents and a selection of 13 sampled cheeses. The prediction error from the hyperspectral image spectra is between that of the two spectrometers. For a typical food sample, the average bias [and replicate standard deviation] was -0.6% [0.5%] for protein and -0.2% [1.3%] for fat. Comparable values for the best spectrometer were -0.2% bias for protein and -0.5% for fat. Some of the advantages of working with hyperspectral images are highlighted: the simultaneous exploration of representations of both spectral and spatial data, and the analysis of concentration profiles and concentration maps all contribute to better characterization of organic and biological materials.

Calibration↗
PubMed1995

Resolution of catecholic tetrahydroisoquinoline enantiomers and the determination of R- and S-salsolinol in biological samples by gas chromatography-mass spectrometry.

Tetrahydroisoquinolines (TIQs) might be formed endogenously and can act centrally to promote a mechanism governing alcohol drinking behaviour. The possibility that biosynthesis occurs through a stereospecific enzymatic reaction is considered. Several TIQs were transformed into diastereomers by a two-step derivation with N-methyl-N-trimethylsilyltrifluoracetamide and R-(-)-2-phenylbutyrylic acid and were analyzed by gas chromatography-mass spectrometry (GC-MS). High resolution of the TIQ enantiomers was achieved. This method was applied to the quantification of the enantiomers of salsolinol (SAL) in urine and plasma of healthy humans. Deuterated SAL was used as the internal standard. SAL was extracted from biological material using phenyl-boronic phase cartridges and transformed into diastereomers. The sensitivity and specificity of the assay permit the determination of the enantiomeric composition of SAL in plasma and urine. The limit of quantification was found to be 100 pg/ml for each enantiomer. The described method has the advantage that optimal resolution of the SAL enantiomers without peak overlapping between analyte and other compounds can be achieved. Contrary to other findings, our GC-MS studies have demonstrated that endogenously formed SAL is racemic in plasma as well as in urine of healthy subjects.

Deuterium↗
PubMed2006

DFTMP, an NMR reagent for assessing the near-neutral pH of biological samples.

A new NMR chemical shift standard and pH indicator, difluorotrimethylsilanylphosphonic acid (DFTMP), is described, and the utility of this reagent is demonstrated for in situ determination of pH in complex biofluids. The pH dependence of this reagent allows accurate in situ determination of aqueous solution pH to within an RMSE of 0.02 pH units over a pH range of 5 to 8. Advantages of this reagent over previously described pH-sensitive components include (1) lack of metal binding affinity, (2) minimal disturbance of endogenous spectral regions, and (3) the potential to function as a multinuclear pH indicator and chemical shift reference point for 19F, 1H, and 31P nuclei. This reagent will be generally useful for NMR experiments on biological systems where the pH needs to be accurately measured at the moment of data acquisition.

Animals↗
PubMed1992

Determination of cadmium in biological samples by inductively coupled plasma atomic emission spectrometry after extraction with 1,5-bis(di-2-pyridylmethylene) thiocarbonohydrazide.

An inductively coupled plasma atomic emission spectrometric method for the determination of trace amounts of cadmium after extraction of the metal into isobutyl methyl ketone containing 1,5-bis(di-2-pyridylmethylene) thiocarbonohydrazide is described. The optimum extraction conditions were evaluated from a critical study of the effects of pH, concentration of extractant, shaking time and ionic strength. The detection limit for cadmium is 0.1 ng ml-1 and the calibration is linear from 0.2 to 140 ng ml-1. The relative standard deviation for ten replicate measurements is 2.9% for 2 ng ml-1 of cadmium. Results from the analysis of some certified biological reference materials are given.

Animals↗
PubMed1990

Evaluation of ibuprofen dimethyl aminoethanol octyl bromide and related active metabolites in biological samples.

AF 150 is ibuprofen dimethyl aminoethanol octyl bromide, endowed with antibacterial and antiinflammatory activities. An indirect HPLC analytical method for AF 150 with UV detection, to be used for pharmacokinetic studies, has been validated in terms of linearity, reproducibility, specificity and sensitivity. Using this method the bioavailability of AF 150 and its active metabolite ibuprofen was investigated following parenteral administration and topical application.

Animals↗
PubMed1997

Study of 4-quinolone antibiotics in biological samples by short-column liquid chromatography coupled with electrospray ionization tandem mass spectrometry.

Simultaneous detection and confirmation of 15 quinolone antibiotics was accomplished by fast short-column liquid chromatography coupled with electrospray ionization tandem mass spectrometry (LC/MS/MS). Several physiochemical parameters such as hydrophobicity and aqueous dissociation constants were calculated from the structural formulas of the quinolone drugs, and their impact on both chromatographic and mass spectrometric behavior was studied. Additionally, a possible influence of bulk solution pH on electrospray detection sensitivity of 4-quinolones was investigated and compared to predictions based on solution-phase equilibria. A signal intensity comparison of the MH+ ions at different pH values for all 15 compounds did not reveal any pH effect, despite variations by several orders of magnitude in equilibrium concentrations in bulk solution. To demonstrate the potential of the LC/MS/MS method, its application to trace analysis in several biological matrices such as milk, salmon, and human urine was investigated. The method was shown to be sensitive with detection limits down to 1 ppb in both milk and salmon tissue. The versatility of the method was also exhibited by utilizing it for rapid identification of urinary metabolites of ciprofloxacin. Finally, a new complementary approach is described for confirmatory analyses of 4-quinolones by means of a quasi-MS/MS/MS technique involving in-source collision-induced dissociation. It is shown that LC/quasi-MS/MS/MS can significantly enhance structural information and, thus, the specificity of analysis for the investigated 4-quinolones.

4-Quinolones↗