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The proposed method determines the activity of cholesterol esterase (CEH) and takes advantage of its ability to catalyze the hydrolysis of cholesterol esters naturally present in human serum. The assay is based on Allain's method of spectrophotometric determination of cholesterol by means of cholesterol oxidase, peroxidase, but using 3,5-dichloro-dihydroxybenzenesulfonic acid (DHBS) as phenolic chromogen and human serum as a source of substrate for the CEH as a novelty. Furthermore, it is characterized by low costs and high precision. It can be employed to control the activity of CE preparations used for the preparation of enzymatic kits for the determination of cholesterol or for screening of potential bacterial enzyme producers.
Dimethoxytritylphosphono-oligonucleotide conjugates have been prepared. They are totally resistant to nucleases present in human serum and do not affect cleavage of a complementary oligoribonucleotide by RNase H. Conjugates possessing a phosphate backbone gave better antisense inhibition of expression of plasminogen activator inhibitor type-1 within endothelial cells as compared with unconjugated oligonucleotides.
Human serum contains several glycosaminoglycans (GAGs), mainly chondroitin sulphates and significantly less of heparan sulphate + heparin and dermatan sulphate. The non-insulin-dependent diabetes mellitus (with vascular complications) was associated with a significant increase in total serum GAG concentration, mainly of chondroitin sulphates and dermatan sulphate, with a simultaneous decrease in heparan sulphate + heparin level. These alterations were much more evident in patients with poor metabolic control. Hyaluronic acid (undetectable in healthy subjects and in patients with good metabolic control) appeared only in trace amounts in poorly controlled diabetic individuals. The obtained data allow to conclude that the diabetes mellitus-associated disturbances in tissue GAG metabolism lead to significant alterations in serum GAG composition.
Calcium and magnesium are known to be necessary for the normal function of various systems in animal and human organisms. There are many diseases caused by abnormal concentration of electrolytes, e.g. arterial hypertension or nervous system diseases such as multiple sclerosis, Mb. Alzheimer or Mb. Parkinson. The mechanisms of homeostasis indicate only the ionized forms of these elements. It is known that ionized calcium serves as an endocellular intermediary in action of enzymes and hormones in cells. Therefore, it is very important to define levels of total and ionized forms of Ca2+ and Mg2+ in blood serum and saliva by the method of atomic absorption spectrometry and to show their diagnostic value for various pathological conditions of a human body. The 39 persons, aged 21 to 47 years take part in these investigations. The results of determinations of calcium and magnesium forms present in human serum and saliva, representing physiological states are presented. The age and daily fluctuations of Ca2+ and Mg2+ content in serum and saliva were studied by atomic absorption spectrometry. The levels of non albumin forms of these elements were found by FAAS. The significance of determination of calcium and magnesium levels in serum and saliva under various pathological conditions (arterial hypertension and osteoporosis) was shown.
Binding affinities of ten polycyclic aromatic hydrocarbons to albumin were determined: anthracene, its eight oxy-derivatives: anthraquinone, 9-anthracenemethanol, 9-anthraldehyde, 9-anthracenecarboxylic acid, 1,4-dihydroxyanthraquinone, 1,5-dihydroxyanthraquinone, 1,8-dihydroxyanthraquinone, 2,6-dihydroxyanthraquinone and benzo[a]pyrene. The quenching of albumin fluorescence was used to measure the PAH — protein interaction. The theoretical curve of calculated fluorescence was fitted to experimental data after necessary corrections regarding PAHs fluorescence and inner filter effect. From the numerical fitting the final association constants were calculated. Anthracene and anthraquinone failed to quench the albumin fluorescence. 9-anthracenecarboxylic acid showed the highest, while 9-anthracenemethanol the weakest albumin binding affinity. The affinity constants determined for 9-anthraldehyde and benzo[a]pyrene were of the same magnitude and indicated low-affinity binding to albumin. The constants obtained for the four dihydroxyanthraquinones were higher, but dissimilar, which suggests that the position of the functional group in anthracene molecule influences the binding constant. Moreover, this study suggests that the type of substituent plays a significant role in PAH-albumin complex formation. The carboxylic group increases the binding affinity of the anthracene molecule the most rather than the presence of both carbonyl and hydroxyl groups. The lowest affinity constants were obtained for aldehyde, methyl and carbonyl substituents.
The sensitivity of bacteria to the bactericidal activity of serum depends on the structure and organization of the bacterial outer membrane. Sialic acid has been found in the O-specific region of bacterial lipopolysaccharide (LPS) and it plays an essential role in protecting Gram-negative bacteria against the bactericidal activity of human and animal serum. The susceptibility of Gram-negative bacilli with sialic acid-containing LPS to the bactericidal action of normal bovine serum (NBS) was determined. The examined strains (Escherichia coli O104 (PCM 270), E. coli O24 (PCM 195), E. coli O56 (PCM 2372), Citrobacter braakii 037 (PCM 2346) and Salmonella entérica ssp. entérica serovar Toucra 048 (PCM 2359) showed variable sensitivity to the bactericidal effect of the serum. The role of the mechanisms of complement activation in the killing process was also determined.
Urinary tract infections are frequently caused by Proteus mirabilis strains. In the previous studies there were defined the complete structures of O-polysaccharide parts of lipopolysaccharides from strains: P. mirabilis O3 (S1959), P. mirabilis O9 and P. mirabilis O18. In the present study it was investigated bactericidal effect of normal human serum (NHS) to P. mirabilis strains. We also focused on the diversity of outer membrane proteins (OMPs) being separated on a gel isolated from tested strains. Serial passage of P. mirabilis O18 in 90% normal bovine serum (NBS) contributed to over-expressing some classes of OMPs.
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