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At least 793 records · Page 44Linked to original sources
PubMed2003

Sample preparation and imaging of erythrocyte cytoskeleton with the atomic force microscopy.

A novel method for the covalent attachment of erythrocytes to glass microscope coverslips that can be used to image intact cells and the cytoplasmic side of the cell membrane with either solid or liquid mode atomic force microscopy (AFM) is described. The strong binding of cells to the glass surface is achieved by the interaction of cell membrane carbohydrates to lectin, which is bound to N-5-azido-2-nitrobenzoyloxysuccinimide (ANBNOS)-coated coverslips (1). The effectiveness of this method is compared with the other commonly used methods of immobilizing intact erythrocytes on glass coverslips for AFM observations. Experimental conditions of AFM imaging of biologic tissue are discussed, and typical topographies of the extracellular and the cytoplasmic surfaces of the plasma membrane in the dry state and in the liquid state are presented. Comparison of the spectrin network of cell age-separated erythrocytes has demonstrated significant loss in the network order in older erythrocytes. The changes are quantitatively described using the pixel height histogram and window size grain analysis.

Cell Adhesion↗
PubMed1988

[Study of profile analysis on urinary free steroids using high performance liquid chromatography with spectrophotometric scanning by photodiode array--application of Girard reagent T for sample preparation].

High performance liquid chromatography (HPLC) has been useful for profile analysis of steroids. However, the conventional extraction of urinary free steroids using urine specimens has some disadvantages because of lots of interfering substances simultaneously extracted from the urine. These substances were usually detected on the chromatogram at the range of relatively short retention time within which some urinary free steroids were heavily contaminated. Therefore it seemed unsuitable for profile analysis of urinary free steroids by HPLC. In this study, we developed a relatively simple and reproducible method for removing the interfering substances by Girard reagent T. In addition, the purity of each extracted free steroids were confirmed by Photodiode Array continuous scanning system, together with 3-dimensional chromatogram as well as contour map analyzed by the attached computer. The extraction procedure was as follows: (1) 10% volume of 24-h specimens of urine included 1 microgram internal standard was charged to Sep-pack C18 cartridge. The cartridge was eluted with 20 micromilligram ethyl acetate and the eluate was evaporated. (2) 10 mg Girard reagent T dissolved in 0.5 micromilligram acetic acid and 0.5 micromilligram ethanol was added to the residue, then left at room temperature for 2 hours. During this time, ketosteroids formed by the action of Girard reagent T turned to be water-soluble hydrazone complex. (3) After the addition of 10 micromilligram cold water, it was adjusted to pH 8 with NaOH and NaHCO3, then washed with 5 volumes of ethyl acetate (non-ketotic fraction). (4) The lower layer were hydrolyzed by adding 0.5 micromilligram concentrated hydrogen chloride and left for an hour at room temperature, then the liberated steroids were extracted with ethyl acetate (ketotic fraction). (5) Ethyl acetate extract was evaporated and redissolved in the mobile phase, then injected to HPLC. To determine the effect of Girard's separation non-ketotic fraction was also applied to HPLC. The results were as follows: (1) 3-dimensional chromatogram and contour map confirmed the effectiveness of our Girard's separation. The chromatogram of ketotic fraction showed the clear separation of the peaks of 4-ene-3-one steroids in ultraviolet absorbance at 246 nm and each steroid peaks were identical to those of the reference steroids. (2) Urinary non-specific substances moving into non-ketotic fraction was complete and satisfactory.(ABSTRACT TRUNCATED AT 400 WORDS)

Adrenal Cortex Hormones↗
PubMed2005

Sample preparation for evaluation of detection limits in X-ray fluorescence spectrometry.

The influence of analyte mass concentration on determination of detection limits in X-ray fluorescence spectrometry has been investigated experimentally. Both the total reflection X-ray fluorescence (TXRF) and the conventional energy-dispersive X-ray fluorescence techniques have been used to derive the dependence of analyte mass concentration on the values of detection limits. Results obtained indicate that values of detection limits are optimum, or in other words, they are closer to the true detection limit of the technique, when analyte concentrations are in the range of 10 times of the detection limit.

Journal Article↗
PubMed2000

Automated sample-preparation technologies in genome sequencing projects.

A robotic workstation system (BioRobot 96OO, QIAGEN) and a 96-well UV spectrophotometer (Spectramax 250, Molecular Devices) were integrated in to the process of high-throughput automated sequencing of double-stranded plasmid DNA templates. An automated 96-well miniprep kit protocol (QIAprep Turbo, QIAGEN) provided high-quality plasmid DNA from shotgun clones. The DNA prepared by this procedure was used to generate more than two mega bases of final sequence data for two genomic projects (Arabidopsis thaliana and Schizosaccharomyces pombe), three thousand expressed sequence tags (ESTs) plus half a mega base of human full-length cDNA clones, and approximately 53,000 single reads for a whole genome shotgun project (Pseudomonas putida).

Arabidopsis↗