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At least 109 records · Page 6Linked to original sources
PubMed2006

Protein chip based miniaturized assay for the simultaneous quantitative monitoring of cancer biomarkers in tissue extracts.

A multiplexed fluorescence immunoassay using a novel planar waveguide technology-based microarray system, ZeptoMARK (Zeptosens), was developed to detect simultaneously urokinase-type plasminogen activator (uPA), plasminogen activator inhibitor-1 (PAI-1), and vascular endothelial growth factor (VEGF) in extracts of breast cancer tissues. The three analytes assay was cross-validated with single-analyte ELISA/chemiluminescence immunosorbent assay tests, revealing good correlations and enhanced assay sensitivities (LODs) of 1 pg/mL for uPA, 33 pg/mL for PAI-1, and 1 pg/mL for VEGF. Values were well within the 80-120% limits for assay recovery and within the +/-20% limits for assay precision. The uPA, PAI-1, and VEGF results obtained from 50 breast cancer cytosols using the protein array system demonstrated that the microarray-based multiplexed assay is a sensitive and robust tool to be used for the simultaneous quantification of cancer markers in small breast cancer tissue samples (core biopsies). The miniaturized, multiplexed assay format has a potential to be used for the quantitative analysis of a larger set of validated markers with significance in disease management.

Biomarkers, Tumor↗
PubMed1999

Simultaneous determination by capillary gas chromatography of organic acids, sugars, and sugar alcohols in plant tissue extracts as their trimethylsilyl derivatives.

A capillary gas chromatographic (GC) method for the simultaneous determination of organic acids, sugars, and sugar alcohols extracted from plant tissues is described. Plant leaves were extracted in 5% (w/v) perchloric acid and neutralized extracts were purified using C18 cartridges. Organic acids, sugars, and sugar alcohols in purified extracts were converted to their trimethylsilyl (TMS)/TMS-oxime derivatives prior to separation and detection by capillary GC with flame ionization detection (FID). Derivatization procedures were investigated in detail and the compounds of interest were readily converted to their TMS/TMS-oxime derivatives using hexamethyldisiazane reagent in acetonitrile solvent (1:6 v/v) at 100 degreesC for 60 min. The derivatives were sufficiently volatile and stable. The FID response to derivatized compounds was generally linear in the concentration range 30-300 microg ml-1, with detection limits in the order of 3-76 ng. The proposed method was demonstrated for the determination of organic acids, sugars, and sugar alcohols in leaf extracts of two native Australian plants.

Biochemistry↗
PubMed1976

Determination of protein in adipose tissue extracts.

An accurate and sensitive method for determination of protein in adipose tissue extracts based on a scaled-down version of the method of Lowry et al. (1951, J. Biol. Chem. 193, 265-275) is described. Protein is quantitatively precipitated by trichloroacetic acid with a nonionic detergent as a bulk-increasing substance and the precipitate is extracted with an organic solvent. This interference caused by lipids, sulfhydryl reagents, and several other compounds is eliminated. The accuracy of the determination of dilute reference protein solutions (2-20 mug/ml) is demonstrated.

Adipose Tissue↗
PubMed1980

The nonspecific inhibitory effect of synovial tissue extracts on leucocyte migration in vitro.

The leucocyte migration inhibition test (LMT) has been used to search for specific antigens in rheumatoid synovial tissue. Synovial samples were collected from 20 patients with rheumatoid arthritis, from 1 patient with ankylosing spondylitis, and from 1 patient with pigmented villonodular synovitis. Inhibitory material was obtained from all 21 synovia with inflammatory disease but not from the noninflammatory synovium. The tissue extracts generally caused nonspecific migration inhibition when tested against a total of 157 pairs of rheumatoid and control leucocytes. However, occasional samples did induce migration inhibition restricted to either rheumatoid or control cells. The inhibitory factor was shown to be membrane associated and of high molecular weight (greater than 10(6) daltons). The results of this extensive study do not support the conclusions drawn from earlier reports based on smaller numbers of experiments. No evidence was obtained for the existence of specific antigenic material associated with the synovial membrane in rheumatoid arthritis.

Adult↗
PubMed2000

Detection of peptide nucleic acids in tissue extracts of treated animals by gel mobility shift assay.

We have developed a sensitive and reproducible gel mobility shift assay to detect PNA oligomers in tissue of treated animals. PNA present in purified tissue extracts of treated animals is hybridized to a 33P-labelled DNA oligomer probe, and analyzed by polyacrylamide gel electrophoresis. The PNA-DNA hybrid migrates more slowly than the DNA probe alone and can be quantified relative to a standard curve. This detection method is useful for detecting PNAs in many different tissues, including brain, heart, kidney, liver, spleen, and serum, as well as cells in culture.

Animals↗
PubMed1997

Determination of serine hydroxymethyltransferase and reduced folate pools in tissue extracts.

Serine hydroxymethyltransferase (SHMT) from all sources tested catalyzes the slow exchange of the pro-2S proton of glycine with solvent protons. In the presence of tetrahydrofolate (H4PteGlun) this exchange rate is increased by about three orders of magnitude. This H4PteGlun-dependent exchange has been developed into a rapid and sensitive assay for both SHMT and H4PteGlun and the one-carbon derivatives of H4PteGlun. The procedure involves incubating [2-3H]glycine, H4PteGlun, and SHMT for 3 min followed by a separation of the exchanged protons in the solvent from the substrate glycine on a small Dowex-50 cation-exchange column at pH 2. In the presence of an excess of H4PteGlun the exchange rate is proportional to nanogram levels of SHMT. In the presence of an excess of SHMT the exchange rate is directly proportional to the concentration of H4PteGlun in the 0.1 to 1 pmol range. The concentration of one-carbon derivatives of H4PteGlun is determined by a preincubation of cell extracts with enzymes that convert each derivative into H4PteGlun. A complete reduced folate pool analysis of a tissue extract can be obtained in less than 2 h once a standard curve has been prepared for H4PteGlun. The method does not distinguish between mono- and polyglutamate forms of the coenzyme.

Animals↗
PubMed2002

A comparison of cell and tissue extraction techniques using high-resolution 1H-NMR spectroscopy.

Analysis of brain metabolites by a wide range of analytical techniques is typically achieved using biochemical extraction methodologies that require either two separate samples or two separate extraction steps to prepare both aqueous and organic metabolite fractions. However there are a number of brain pathologies in which both aqueous metabolite and lipid changes occur so that a simultaneous extraction of both fractions would be valuable. The methanol-chloroform (M/C) technique enables extraction of both aqueous metabolites and lipids simultaneously. It is already well established for lipid extraction of cells and tissue but its efficiency and reproducibility for extraction of aqueous metabolites is unknown. Therefore, we compared the aqueous metabolite yield and the reproducibility of the M/C method to the commonly used perchloric acid (PCA) method, using 1H-NMR spectroscopy of adult rat brain and purified rat astrocyte culture extracts. The results indicate that M/C is a superior technique for aqueous metabolite extraction from both brain tissue and cells when compared to the PCA method. The M/C extraction technique enables the simultaneous extraction of both lipids and aqueous metabolites from a single sample using small solvent-volumes, making it well suited for NMR investigations of both tissues and cells.

Animals↗
PubMed2005

Comparisons of brain metabolites observed by HRMAS 1H NMR of intact tissue and solution 1H NMR of tissue extracts in SIV-infected macaques.

The objective of this study was to compare ex vivo proton high-resolution magic angle spinning magnetic resonance spectra of intact tissue with those spectra obtained by solution (1)H NMR of brain extracts of the same sample. Sixteen brain tissue samples from simian immunodeficiency virus-infected rhesus macaques from both frontal cortex and putamen were evaluated by comparing brain metabolite quantities of N-acetylaspartate (NAA), choline-containing compounds (Cho), myo-inositol (MI), creatine (Cr), lactate (Lac), glutamate (Glu) and acetate (Ace). The ratios of the individual NMR peak areas of all metabolites relative to the creatine peak area were calculated. Linear regression analysis revealed significant correlations between measurements using the two methods. The strength of the correlations varied depending on the metabolite studied. We found highly significant correlations for NAA/Cr (r2 = 0.77; p < 0.0001), NAA + Ace/Cr (r2 = 0.73; p < 0.0001) and MI/Cr (r2 = 0.75; p < 0.0001). We observed somewhat less strong correlations for Glu/Cr (r2 = 0.54; p < 0.002) and Lac/Cr (r2 = 0.54; p < 0.002). There was a substantially weaker correlation for Cho/Cr (r2 = 0.32; p = 0.02). When plotting the metabolite ratios obtained by 1H HRMAS NMR of the intact tissue sample on the ordinate vs 1H NMR of the tissue extract on the abscissa, most metabolites exhibited a slope close to unity, and a positive intercept probably due to macromolecular contributions to the MAS spectra. The slope for Cho/Cr was substantially less than unity. Generally, samples from the frontal cortex showed a better correlation between intact and extracted tissue samples than putamen. This is most prominent in the cases of NAA/Cr and Cho/Cr. We conclude that both methods provide substantially the same information for most major brain metabolites, with the exception of the Cho resonance.

Animals↗
PubMed1987

Immunoreactive neuromedin B and neuromedin C: distribution and molecular heterogeneity in rat and human tissue extracts.

Immunoreactive neuromedin B and neuromedin C were characterized and measured in rat and human tissue extracts, using radioimmunoassay combined with gel filtration and high performance liquid chromatography in order to clarify tissue distribution and molecular structure. There are two distinct systems of bombesin-like peptide found in rat and human tissue. The neuromedin B family, including neuromedin B and big neuromedin B, is a major bombesin-like peptide in the brain; the neuromedin C gastrin-releasing peptide family is found mainly in the alimentary tract. Neuromedin B is the major and big neuromedin B the minor constituent of central nervous neuromedin B immunoreactivity. Gastrointestinal neuromedin C immunoreactivity is composed chiefly of neuromedin C in rats, and two N-terminally extended forms of neuromedin C including gastrin-releasing peptide in man. It is probable that molecular heterogeneity reflects species variation.

Animals↗
PubMed2005

[Possibilities of investigation of the tissues extracts using method of the laser correlative spectroscopy in differential diagnosis of the thyroid gland cancer and chronic autoimmune thyroiditis].

The results of investigation of the intraoperatively excised tissues extracts using method of the laser correlative spectroscopy were analyzed. Application of laser correlative spectroscopy permit to differentiate the thyroid gland cancer and chronic autoimmune thyroiditis in 82% of observations.

Chronic Disease↗
PubMedDate not supplied

Fluorometric estimation of taurine in tissue extracts and biological fluids.

A fluorometric technique was developed to estimate taurine in tissue extracts and body fluids. Dowex-AG and Biorad-AG columns were used to separate this amino acid from other components. Interference by Glycerophosphoryl ethanolamine was removed by hydrolysis of the sample with 6N HCl. The fluorogen used was fluorescamine.

Animals↗
PubMed1983

Estimation of the blood contamination of tissue extracts.

A method based on measurement of absorbance at 540 or 580 nm is described. The results obtained were closely correlated to the actual level of blood present in different tissues, as determined by use of radiolabeled albumin. The present technique is much simpler than earlier methods used to determine the degree of blood contamination of tissue extracts. Hence, it may find wide application in the life sciences.

Animals↗
PubMed1992

Improved and simplified tissue extraction method for quantitating long-chain acyl-coenzyme A thioesters with picomolar detection using high-performance liquid chromatography.

A method has been developed that permits rapid and easy tissue extraction of long-chain acyl-coenzyme A (acyl-CoA) thioesters with sensitive quantitation by reversed-phase high-performance liquid chromatography (RP-HPLC). Tissue homogenants are extracted using a reserve Bligh-Dyer technique, and long-chain acyl-CoA esters are harvested in the methanolic aqueous phase. Complex lipids and phospholipids are removed in the chloroform-rich organic Bligh-Dyer second phase, and long-chain acyl-CoA compounds are further purified from the methanolic aqueous Bligh-Dyer first phase on C18 extraction columns after removal of the methanol. The eluted and purified acyl-CoA esters are then quantitated by RP-HPLC using heptadecanoyl-CoA as an internal standard resulting in a detector sensitivity of about 12 pmol. Ten long-chain acyl-CoA esters from C12:0 to C20:4 were identified and separated from canine renal cortex and murine liver samples. The predominant acyl-CoA peaks from both kidney and liver were 14:0, 16:1, 16:0, 18:1, 18:2 and 20:4. Murine liver also produced 18:0 and all peaks disappeared after alkaline hydrolysis of the samples. This extraction and quantitation technique can successfully be used for tissue samples as small as 20 mg, and many samples can be processed in a short period of time. The simplicity of the extraction procedure and the sensitivity of the assay make this an attractive alternative approach to quantitating long-chain acyl-CoA thioesters from complex biological samples such as tissues.

Acyl Coenzyme A↗
PubMed2004

Casuarina cunninghamiana tissue extracts stimulate the growth of Frankia and differentially alter the growth of other soil microorganisms.

Aqueous extracts of host plant Casuarina cunninghamiana tissue altered the in vitro growth of its diazotrophic microsymbiont Frankia and a selection of other soil microorganisms. The growth of actinomycetous Frankia strains, 55005. AvcI1, CesI5, CjI82 001, and Cj was stimulated by aqueous extracts of C. cunninghamiana tissue. Green cladodes (photosynthetic branches), unsuberized roots, and suberized roots were more stimulatory than dry cladodes and seed tissue. Aqueous extracts of green cladodes of C. cunninghamiana most stimulated the growth of Casuarina-derived Frankia strains CjI82 001 and 55005. The growth of isolates of soil bacteria Bradyrhizobium japonicum, Arthrobacter globiformis and Bacillus subtillis and of the soil fungi Penicillium oxalicum and Arthroderma cookiellum was either inhibited or not affected by cladode extracts. Cladode extracts stimulated the growth of the actinomycete Streptomyces albus and the fungus Rhizopus homothallicus. The magnitude (as great as 100%) of the increase in growth caused by tissue extracts for the Casuarina-derived Frankia strains relative to other soil microbes suggests a host-specific enhancement of the microsymbiont.

Bacteria↗
PubMed1979

Morphologic study of the action of lymphoid tissue extracts in peroxidase arthritis.

By means of modelling arthritis by horseradish peroxidase it was possible to reproduce a process corresponding to immune synovitis. A considerable reduction of immunomorphological phenomena in joints, lymphoid tissue, heart, liver and kidneys was favoured by application of lymphoid tissue extracts. Comparison between the extracts of spleen and lymph nodes showed a more pronounced effect of spleen extract.

Animals↗