| 1. |
- Nahálková, Jarmila, et al.
(författare)
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Affinity analysis of lectin interaction with immobilized C- and O-gylcosides studied by surface plasmon resonance assay
- 2002
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Ingår i: Journal of Biochemical and Biophysical Methods. - 0165-022X .- 1872-857X. ; 52:1, s. 11-18
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Tidskriftsartikel (refereegranskat)abstract
- A biosensor based on the surface plasmon resonance (SPR) principle was used for kinetic analysis of lectin interactions with different immobilized saccharide structures. A novel affinity ligands beta-D-glycopyranosylmethylamines derived from common D-aldohexoses linked to the carboxymethyl dextran layer of the SPR sensor surface served for interactions with a wide range of lectins. The method of preparation and use of the beta-D-mannopyranosyl glycosylated sensor surface was described. The results of affinity analysis of lectin-ligand interactions were evaluated and compared with data obtained from measurements using commercially available p-aminophenyl alpha-D-glycopyranosides. Possible applications and advantages of C- and O-glycosylated SPR biosensors are discussed.
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| 2. |
- Haupt, Dan, et al.
(författare)
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Separation of (R)- and (S)-naproxen using micellar chromatography and an alpha 1-acid-glycoprotein column : application for chiral monitoring in human liver microsomes by coupled-column chromatography
- 1992
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Ingår i: Journal of Biochemical and Biophysical Methods. - 0165-022X .- 1872-857X. ; 25:4, s. 273-284
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Tidskriftsartikel (refereegranskat)abstract
- A column-switching system for fast determination of (R)- and (S)-naproxen in liver microsomes has been developed. The centrifuged sample was injected directly onto a pre-column with octadecylcoated silica. The retained analytes were then directed to an alpha 1-AGP column using a mobile phase composed of phosphate buffer (pH 6.5), dimethylocytylamine (30 mM) and the nonionic surfactant, Tween 20 (40 g/l). The method gave high absolute recoveries and good repeatabilities: 99.6% (1.7% relative standard deviation) and 94.9% (2.4% R.S.D.) for the (R)- and (S)-naproxen, respectively. The use of a surfactant in combination with an aliphatic amine in the mobile phase involves reduced retention times with retained enantioselectivity. Furthermore, the presence of the surfactant makes it possible to inject biological samples directly into the chromatographic system.
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| 3. |
- Lagerquist Hägglund, Christine, et al.
(författare)
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Centrifugal and chromatographic analyses of tryptophan and tyrosine uptake by red blood cells and GLUT1 proteoliposomes with permeability estimates and observations on dihydrocytochalasin B
- 2003
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Ingår i: Journal of Biochemical and Biophysical Methods. - 0165-022X .- 1872-857X. ; 55:2, s. 127-140
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Tidskriftsartikel (refereegranskat)abstract
- We analyzed transport into liposomes and proteoliposomes, separated the free and internalized radioactively labeled substrates by size-exclusion chromatography (SEC) and observed a net influx owing to nonfacilitated diffusion across the lipid bilayers during the separation. The permeabilities (10(-9) cm/s) of glucose transporter (GLUT1) proteoliposomes were estimated to be 4.6, 1.0, 1.4 and 2.1 for D-glucose, L-glucose, L-Tyr and L-Trp, respectively; 15, 3.3, 5.1 and 2.1 times higher than the corresponding permeabilities of liposomes. These values indicated that GLUT1 did not transport Tyr or Trp, or transported Tyr, and only Tyr, slowly. This interpretation was supported by further analyses. Dihydrocytochalasin B inhibited the transport of Tyr and, partially, Trp into human red blood cells (centrifugal analyses). It did not inhibit Tyr and Trp influx into GLUT1 proteoliposomes, but partitioned strongly into the bilayers and seemed to make them fragile. The GLUT1 inhibitor cytochalasin B and the GLUT1 substrate 2-deoxy-D-glucose did not inhibit Tyr transport into the cells. Upon immobilized biomembrane affinity chromatography, Trp decreased the cytochalasin B retardation by GLUT1 only at levels far above the physiological Trp concentration. Ethanol (commonly added to aqueous solutions for enhancing a compound's solubility) halved the retardation at 4% (v/v) concentration. Drastic modification of the SEC method is required to allow permeability measurements with nonlabeled and highly permeable substrates.
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| 4. |
- Nordstrom, T., et al.
(författare)
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Method for one-step preparation of double-stranded DNA template applicable for use with Pyrosequencing (TM) technology
- 2002
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Ingår i: Journal of Biochemical and Biophysical Methods. - 0165-022X .- 1872-857X. ; 52:2, s. 71-82
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Tidskriftsartikel (refereegranskat)abstract
- A new one-step method for fast and efficient preparation of double-stranded DNA template, suitable for use with Pyrosequencing(TM) technology, has been developed. In the new method, two different types of oligonucleotides were used to prevent reannealing of remaining PCR primers to the template: oligonucleotides complementary to the PCR primers and 3'-end modified oligonucleotides with the same sequence as the PCR primers. Advantages with the new strategy are: (i) faster and simpler template preparation procedure (one-step); (ii) no need for exonuclease I treatment; and (iii) less problem with unspecific priming from loop structures and dimers. By careful oligonucleotide design, and/or by addition of single-stranded DNA-binding protein, problems with unspecific sequence signals due to mispriming can be reduced. The new method was used for analysis of genotype variations within the renin-angiotensin-aldosterone system.
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| 5. |
- Södergren, E, et al.
(författare)
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Re-evaluation of the ferrous oxidation in xylenol orange assay for the measurement of plasma lipid hydroperoxides.
- 1998
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Ingår i: Journal of Biochemical and Biophysical Methods. - : Elsevier BV. - 0165-022X .- 1872-857X. ; 37:3, s. 137-46
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Tidskriftsartikel (refereegranskat)abstract
- The ferrous oxidation in xylenol orange version 2 (FOX2) assay coupled with triphenylphosphine has recently been employed for the measurement of total plasma hydroperoxides (ROOHs). In this study, we have evaluated sample handling and the effect of storage conditions on ROOH levels in human plasma (n = 32). Mean level of ROOHs in fresh plasma was 8.35 +/- 3.09 mumol/l (range 4.03-19.5 mumol/l). Addition of butylated hydroxytoluene (BHT) immediately after sample collection had no effect on the concentration of ROOHs. Storage of samples at -70 degrees C for 6 weeks was associated with a variable degree of loss of detectable ROOHs. A mean ROOH level of 6.00 +/- 2.23 mumol/l (range 2.88-13.5 mumol/l) was recorded after 6 weeks of storage at -70 degrees C. There was no difference in the mean level of ROOHs between samples stored for 6 and 60 weeks at -70 degrees C. Inclusion of BHT had no effect on the stability of plasma ROOHs during prolonged storage. Intra-assay coefficients of variation were < 12%, with the lowest variation in fresh samples (7.6%). In conclusion, these results suggest that the FOX2 assay should be a useful tool for measurement of ROOHs in fresh plasma samples but not in stored samples.
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| 6. |
- Eriksson, Håkan
(författare)
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Controlled release of preservatives using dealuminated zeolite Y
- 2008
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Ingår i: Journal of Biochemical and Biophysical Methods. - : Elsevier BV. - 0165-022X .- 1872-857X. ; 70:6, s. 1139-1144
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Tidskriftsartikel (refereegranskat)abstract
- This study demonstrates that dealuminated zeolite Y can act as a depot after adsorption of phenol derived preservatives. Upon suspension of zeolite loaded with the preservative m-cresol, equilibrium was quickly reached between free and adsorbed m-cresol. The equilibrium concentration of m-cresol was below 1 mM, however, it was enough to kill bacteria such as Escherichia coli and Staphylococcus aureus under metabolically active conditions. Killing of bacteria were not obtained under non-proliferating conditions and m-cresol was only released from the zeolite upon bacterial activity. Together, these results demonstrate an interesting potential use of dealuminated zeolite Y containing adsorbed preservatives for preventing microbial growth in numerous applications in industry and clinical setting.
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| 7. |
- Hakkarainen, Minna
(författare)
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Developments in multiple headspace extraction
- 2007
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Ingår i: Journal of Biochemical and Biophysical Methods. - : Elsevier BV. - 0165-022X .- 1872-857X. ; 70:2, s. 229-233
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Forskningsöversikt (refereegranskat)abstract
- This paper reviews new developments in multiple headspace extraction (MHE), especially its combination with two miniaturized extraction techniques, solid-phase microextraction (SPME) and single-drop microextraction (SDME). The combination of the techniques broadens the applicability of SPME and SDME to quantitative determination of analytes in complex liquid and solid matrixes. These new methods offer several advantages over traditional liquid-solid, liquid-liquid and headspace extraction techniques. The potential applications include extraction of volatiles and semivolatiles from environmental and physiological samples and from different polymer products such as medical and biomedical materials, food packaging and building materials. The theoretical principals of the techniques are also briefly reviewed.
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| 8. |
- Hedin, Eva M. K., et al.
(författare)
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Low microwave-amplitude ESR spectroscopy : Measuring spin-relaxation interactions of moderately immobilized spin labels in proteins
- 2004
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Ingår i: Journal of Biochemical and Biophysical Methods. - : Elsevier BV. - 0165-022X .- 1872-857X. ; 60:2, s. 117-138
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Tidskriftsartikel (refereegranskat)abstract
- Electron spin resonance (ESR) spectroscopy in combination with site-directed spin labeling (SDSL) is a powerful tool for determining protein structure, dynamics and interactions. We report here a method for determining interactions between spin labels and paramagnetic relaxation agents, which is performed under subsaturating conditions. The low microwave-field amplitude employed (h(1) < 0.36 G) only requires standard, commercially available ESR equipment. The effect of relaxation enhancement on the spin-spin-relaxation time, T-2e, is measured by this method, and compared to classical progressive power saturation performed on a free spin label, (1-oxyl-2,2,5,5-tetramethyl-Delta(3)-pyrroline-3-methyl)methanethiosulfonate (MTSL), and a spin-labeled protein (Thermomyces lanuginosa lipase, TLL-1252C), employing the water-soluble relaxation agent chromium(III) oxalate (Crox) in concentrations between 0-10 mM. The low-amplitude theory showed excellent agreement with that of classical power saturation in quantifying Crox-induced relaxation enhancement. Low-amplitude measurements were then performed using a standard resonator, with Crox, on 11 spin-labeled TLL mutants displaying rotational correlation times in the motional narrowing regime. All spin-labeled proteins exhibited significant changes in T-2e. We postulate that this novel method is especially suitable for studying moderately immobilized spin labels, such as those positioned at exposed sites in a protein. This method should prove useful for research groups with access to any ESR instrumentation.
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| 9. |
- Hjerten, Maria, et al.
(författare)
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Renewable enzyme reactors based on beds of artificial gel antibodies.
- 2008
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Ingår i: Journal of Biochemical and Biophysical Methods. - : Elsevier BV. - 0165-022X .- 1872-857X. ; 70:6, s. 1188-1191
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Tidskriftsartikel (refereegranskat)abstract
- A novel approach is described for the synthesis of beds for enzyme reactors. The method is based on the use of artificial antibodies in the form of polyacrylamide gel particles with diameters around 0.1-0.3 mm. These gel particles mimic protein antibodies, raised in experimental animals, in the sense that they selectively recognize and adsorb only the protein present during the preparation of the "antibodies". The gel antibodies have several advantages over conventional protein antibodies, which can be taken advantage of in the design of enzyme reactors; for instance, if upon prolonged use the immobilized enzyme loses its activity it can easily be replaced by an active enzyme, which is not possible when the enzyme is immobilized via a conventional protein antibody (a new bed with immobilized protein antibodies must be prepared); and equally or more remarkable: the enzyme can be applied in the form of a non-purified extract since the selectivity of the artificial gel antibodies is so high that they will "fish-out" the enzyme, but no other proteins in the extract. In addition, no preconcentration of the enzyme solution is required prior to the immobilization, since the enzyme is enriched at the top of the column upon the application. These unique properties make enzyme reactors based on artificial gel antibodies very attractive, also in process chromatography. The potential application range of the artificial gel antibodies is enormous since the same method for their synthesis can be used independent of the structure and the size of the "antigen"; for instance, renewable biosensors based on gel antibodies for the selective detection of protein biomarkers, as well as pathogenic viruses, bacteria, and spores (for instance Anthrax) should not be difficult to design.
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| 10. |
- Kalbin, Georgi, et al.
(författare)
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Effects of UV-B in biological and chemical systems : equipment for wavelength dependence determination
- 2005
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Ingår i: Journal of Biochemical and Biophysical Methods. - : Elsevier BV. - 0165-022X .- 1872-857X. ; 65:1, s. 1-12
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Tidskriftsartikel (refereegranskat)abstract
- The thinning of the stratospheric ozone layer has prompted a large number of studies of UV-B-induced effects in biological and chemical systems. The wavelength dependency of such effects is of interest from mechanistic, physiological or economic points of view. Here, we describe an apparatus for determining the wavelength dependency of UV-B effects in biological and chemical systems. The apparatus consists of a high intensity UV radiation source and narrow bandpass filters to produce UV radiation in even intervals (between 280 and 360 nm). The usefulness of the equipment is demonstrated in two different systems: 1) Chalcone synthase (CHS) gene is up-regulated by UV-B radiation. Therefore quantitative analysis of the CHS gene expression was chosen in the present investigation for studies of the wavelength dependency of gene expression regulation in plants. Maximum induction of CHS expression was found at 300 nm with a 12-fold induction compared with the control; 2) The wavelength dependency of formation of dioxin-like photoproducts from the brominated flame retardant decabrominated diphenyl ether (DeBDE) is described. This is an example of UV-B-induced conversion of non-toxic species into a number of products of which some may be toxic in the environment. In the UV interval studied, the highest dioxin-like activity was found in the sample irradiated at 330 nm and therefore this wavelength is most important for the mechanism involved in photoconversion of DeBDE.
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