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J Nilsson

Publications and source records attributed to J Nilsson.

At least 109 records · Page 6Linked to original sources
PubMed1999

Homogeneous sample preparation for automated high throughput analysis with matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry.

This work presents a simple method for obtaining homogeneous sample surfaces in matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOFMS) for the automated analysis of peptides and proteins. The sample preparation method is based on applying the sample/matrix mixture onto a pre-deposited highly diluted matrix spot. The pre-deposited crystals act as seeds for the new sample containing crystals which become much smaller in size and more evenly distributed than with conventional methods. This 'seed-layer' method was developed, optimised and compared with the dried-droplet method using peptides and proteins in the 1000-20,000 Da range. The seed-layer method increases the surface homogeneity, spot to spot reproducibility and sample washability as compared with the commonly used dried-droplet method. This methodology is applicable to alpha-cyanohydroxycinnamic acid, sinapinic acid and ferulic acid, which all form homogeneous crystal surfaces. Within-spot variation and between-spot variation was investigated using statistics at a 95% confidence level (n = 36). The statistical values were generated from more than 5000 data points collected from 500 spectra. More than 90% of the sample locations results in high intensity spectra with relatively low standard deviations (RSDs). Typically obtained data showed an RSD of 19-35% within a sample spot as well as in-between spots for proteins, and an RSD of < or = 50% for peptides. Linear calibration curves were obtained within one order of magnitude using internal calibration with a point-RSD of 3% (n = 10). The sample homogeneity allows mass spectra (average of 16 laser shots) to be obtained on each individual sample within 15 sec, whereby a 100 spot target plate can be run in 25 min. High density target plates using the seed-layer method were prepared by spotting approximately 100 picoliter droplets onto the target, resulting in sample spots < or = 500 microns in diameter using a flow-through piezo-electric micro-dispenser. By using this automated sample preparation step lower standard deviations are obtained in comparison to manually prepared samples.

Coumaric Acids↗
PubMed1999

Production, purification, and luminometric analysis of recombinant Saccharomyces cerevisiae MET3 adenosine triphosphate sulfurylase expressed in Escherichia coli.

ATP sulfurylase cDNA from MET3 on chromosome X of Saccharomyces cerevisiae was amplified and cloned, and recombinant ATP sulfurylase was expressed in Escherichia coli. The synthesis of ATP sulfurylase was directed by an expression system that employs the regulatory genes of the luminous bacterium Vibrio fischeri. A soluble, biologically active form was purified to electrophoretic homogeneity from lysates of recombinant E. coli by ammonium sulfate fractionation, ion-exchange chromatography, and gel filtration. The specific activity of the purified enzyme was estimated to 140 U/mg. The apparent molecular mass of the recombinant enzyme was determined by gel filtration to be 470 kDa, which indicates that the active enzyme is an octamer of identical subunits (the molecular mass of a single subunit is 59.3 kDa). The ATP sulfurylase activity was monitored in real time by a very sensitive bioluminometric method.

Chromatography, Gel↗
PubMed1999

Designed four-helix bundle catalysts--the engineering of reactive sites for hydrolysis and transesterification reactions of p-nitrophenyl esters.

Four-helix bundle proteins have been designed that catalyze the hydrolysis and transesterification reactions of p-nitrophenyl esters by a cooperative nucleophilic and general acid mechanism. The catalysts consist of two 42-residue peptides that fold into helix-loop-helix motifs and dimerise. They have previously been shown to recognize anionic and hydrophobic substrates and to follow saturation kinetics. The catalytic entity is a HisH(+)-His pair in a helical segment spaced i, i+4, which can be supplemented by arginines and lysines in the adjacent helix. The binding residues have now been optimized for the catalysis of mono-p-nitrophenyl fumarate hydrolysis and found to vary with the location of the site. The catalytic efficiency of the HisH(+)-His site in helix II in positions 30 and 34 is enhanced by the introduction of arginine and or lysine residues in positions 11 and 15, but not in 8 and 11 or in 15 and 19. The most efficient catalyst using this site, JNIIR11K15, catalyses the reaction with a second-order rate constant of 0.134 M(-1) s(-1) in aqueous solution at pH 5.1 and 290 K. The second-order rate constant is larger than those of the corresponding sites with 'longer' and 'shorter' binding residues. Similar experiments have shown that the efficiency and selectivity of catalysts based on a HisH(+)-11-His-15 site in helix I are enhanced the most by the introduction of Lys-30 and Arg-34.

Amino Acid Sequence↗
PubMed1999

Multivariate quantitative structure-pharmacokinetic relationships (QSPKR) analysis of adenosine A1 receptor agonists in rat.

The aim of this study was to investigate the feasibility of a quantitative structure-pharmacokinetic relationships (QSPKR) method based on contemporary three-dimensional (3D) molecular characterization and multivariate statistical analysis. For this purpose, the programs SYBYL/CoMFA, GRID, and Pallas, in combination with the multivariate statistical technique principal component analysis were employed to generate a total of 16 descriptor variables for a series of 12 structurally related adenosine A1 receptor agonists. Subsequently, the multivariate regression method, partial least squares, was used to predict clearance (CL), volume of distribution (VdSS) and protein binding (fraction unbound, fU). The QSPKR models obtained could account for most of the variation in CL, VdSS, and fU (R2 = 0.82, 0.61 and 0.78, respectively). Cross-validation confirmed the predictive ability of the models (Q2 = 0.59, 0.41 and 0.62 for CL, VdSS, and fU, respectively). In conclusion, we have developed a multivariate 3D QSPKR model that could adequately predict overall pharmacokinetic behavior of adenosine A1 receptor agonists in rat. This methodology can also be used for other classes of compounds and may facilitate the further integration of QSPKR in drug discovery and preclinical development.

Animals↗
PubMed1999

Prostate cancer registration in four Swedish regions 1996-- differences in incidence, age structure and management.

INTRODUCTION: In 1996 registration of prostate cancer in four of the six Swedish regions was started to facilitate evaluation of geographical variations in incidence and treatment. MATERIAL AND METHODS: For all cases of prostate cancer, personal identification number, tumour stage, tumour grade and primary treatment were registered. RESULTS: In the four regions covered by the register, 3541 cases of prostate cancer were registered. Altogether there were 5795 cases of prostate cancer diagnosed in Sweden the same year. The age-standardized incidence varied from 89/100000 to 169/100000 among counties. The proportion of localized tumours correlated positively to the incidence (p < 0.05) and negatively to mean age at diagnosis (p < 0.01). There was also a significant positive correlation between the proportion of localized tumours and the percentage of patients given curative treatment. All registered variables showed large geographical variations, especially concerning percentage of T1c tumours, treatment of localized tumours and choice of palliative treatment. CONCLUSION: Diagnostic activity varied considerably among counties, resulting in large variation in age-standardized incidence. High incidence is associated with a larger proportion of localized tumours, which, in turn, is associated with early age at diagnosis. In counties where a policy of detecting tumours early is practised, curative treatment is also given more often. Treatment of localized tumours and preference for palliative treatment seem to depend on local traditions. The lack of cytological and histopathological standards makes geographical comparisons based on tumour grade impossible.

Age Distribution↗
PubMed1999

A common functional polymorphism (C-->A substitution at position -863) in the promoter region of the tumour necrosis factor-alpha (TNF-alpha) gene associated with reduced circulating levels of TNF-alpha.

Tumour necrosis factor-alpha (TNF-alpha) plays a key role in orchestrating the complex events involved in inflammation and immunity. Accordingly, TNF-alpha has been implicated in a wide range of autoimmune and infectious diseases, but also in conditions such as obesity and insulin resistance. The regulation of TNF-alpha expression in man is indicated to be partly genetically determined. We therefore screened a 1263 bp section of the proximal promoter of the TNF-alpha gene for common genetic variants affecting the transcriptional activity of the gene. Here we report the characterization of a common functional polymorphism in the promoter region of the TNF-alpha gene, a C-->A substitution at position -863. Electromobility shift assays provided evidence for a distinct difference in the binding of monocytic and hepatic nuclear factors to the -863C and -863A alleles. The rare -863A allele was associated with 31% lower transcriptional activity ( P < 0.001) in chloramphenicol acetyltransferase (CAT) reporter gene studies in human hepatoblastoma (HepG2) cells, indicating that the-863C/A polymorphism influences the basal rate of transcription of the TNF-alpha gene in vitro. Allele frequencies were 0.83/0.17 amongst 254 apparently healthy men of Swedish origin, aged 35-50 years. In 156 men, the -863C/A polymorphism was associated with the serum TNF-alpha concentration, carriers of the rare A allele having a significantly lower TNF-alpha level ( P < 0.05). It is concluded that the common-863C/A polymorphism in the promoter region of the TNF-alpha gene is functional in vitro in monocytic and hepatic cells and influences the serum TNF-alpha concentration in vivo in healthy middle-aged men.

Adult↗
PubMed1999

Expression of the T-cell markers CD3, CD4 and CD8 in healthy and atopic children during the first 18 months of life.

BACKGROUND: There is little information available about the development of T-cell immunity in healthy and atopic children. We have studied prospectively the mean fluorescence intensity of the T-cell receptor complex-associated CD3, CD4 and CD8 in relation to atopic family history (AFH) and the development of atopic disease. METHODS: Children with a defined AFH (n = 172) were followed from birth to 18 months and the cumulative history of atopic disease was recorded. Blood samples were obtained at birth and at 18 months, and in a subgroup of 78 children also at 3, 6 and 12 months. Multicolour flow cytometry was used to analyse pan T-cells (CD3+CD45+CD14-), T-helper-(CD3+CD4+) and T-cytotoxic-(CD3+CD8+) cells. RESULTS: At 18 months, 31 children were atopic and 118 non-atopic. Children who developed atopic disease had a higher CD4 expression (mean fluorescence intensity, MFI) on CD4+CD3+ lymphocytes at birth and at 3 months, particularly as compared with non-atopic children without AFH. Furthermore, the CD3 expression on CD3+CD45+CD14- lymphocytes increased more slowly with age in children with double atopic heredity, as compared with children with no or only one atopic family member. CONCLUSIONS: The higher expression of the CD4 receptor in early infancy in children who developed atopic disease compared with non-atopics suggests a delayed expression in T-helper cells. Children with a strong AFH had a slower increase in the expression of CD3, indicating a delayed T-cell maturation.

CD3 Complex↗
PubMed1999

Oxidized LDL and lysophosphatidylcholine stimulate plasminogen activator inhibitor-1 expression in vascular smooth muscle cells.

Plasminogen activator inhibitor-1 (PAI-1) functions as an important regulator of fibrinolysis by inhibiting both tissue-type and urokinase-type plasminogen activator. PAI-1 is produced by smooth muscle cells (SMCs) in atherosclerotic arteries, but the mechanisms responsible for induction of PAI-1 in SMCs are less well understood. In cultured human aortic SMCs, PAI-1 mRNA expression and protein secretion were increased after incubation with oxidized low-density lipoprotein (LDL) and the lipid peroxidation product lysophosphatidylcholine, whereas the effects of native LDL on PAI-1 production and release were more variable and did not reach statistical significance. The effect of LDL on arterial expression of PAI-1 in vivo was also studied in an animal model. Intravenous injection of human LDL in Sprague-Dawley rats resulted in accumulation of apolipoprotein B in the aorta within 12 hours as assessed by immunohistochemical testing. Epitopes specific for oxidized LDL began to develop in the aorta 12 hours after injection of LDL and peaked at 24 hours; this peak was accompanied by intense expression of PAI-1 immunoreactivity in the media. Also, increased aortic expression of PAI-1 mRNA after LDL injection was detected by using in situ hybridization. The transcription factor activator protein-1, which is known to bind to the promoter of the PAI-1 gene, was activated in the aortic wall 24 hours after LDL injection as assessed by electrophoretic mobility shift assay. Pretreatment of LDL with the antioxidant probucol decreased expression of oxidized LDL and PAI-1 immunoreactivity and activator protein-1 induction in the aorta but did not affect expression of apolipoprotein B immunoreactivity. These findings demonstrate that LDL oxidation enhances secretion of PAI-1 from cultured SMCs and that a similar mechanism may be involved in vascular expression of PAI-1.

Animals↗
PubMed1999

Presence of oxidized low density lipoprotein in nonrheumatic stenotic aortic valves.

The aim of the present study was to analyze if LDL particles trapped in stenotic aortic valve tissue undergo oxidative modification. Degenerative aortic stenosis affects >3% of the population >75 years of age in the Western world. Recent studies have revealed the presence of a chronic inflammatory process similar to what has been described in other degenerative diseases such as atherosclerosis. However, the underlying disease mechanisms of degenerative aortic stenosis still remain largely unknown. Six tricuspid stenotic valves, obtained at valve replacement, were compared with 3 control valves collected from hearts taken out during transplantation. The stenotic valves and the control valves were examined by immunohistochemistry, using antibodies against apoB, 4-hydroxynonenal-modified LDL, leukocytes, and HLA-DR. All valves were also stained with oil red O for neutral lipids. Extracellular neutral lipids were found in all stenotic valves, extending from the bases along the fibrosa layer. This lipid colocalized with apoB- and 4-hydroxynonenal-modified LDL immunoreactivity. 4-Hydroxynonenal-modified LDLs were present around calcium deposits, subendothelially, and in the deeper layer of the fibrosa. There was also a colocalization with macrophages, T lymphocytes, and HLA-DR expression. Control valves had a thin area of neutral lipid accumulation, a small amount of apoB, but no signs of inflammation. A distinct colocalization between oxidized LDLs, T-lymphocyte accumulation, and calcium deposits suggests that oxidized lipids may play a role in the disease process.

Adult↗
PubMed1999

Phosphatase activity in the arterial wall after balloon injury: effect of somatostatin analog octreotide.

Phosphorylation of transcription factors fos/jun dimer activator protein (AP)-1 and nuclear factor-kappaB (NF-kappaB) plays a cardinal role in vascular smooth muscle cell (SMC) response to growth stimuli. Activity of protein tyrosine (PTP) and serine/threonine phosphatases (PP2A, B, and C) regulates in balance with the activity of protein kinases the level of transcription factor phosphorylation. Somatostatin analog octreotide stimulates phosphatase activity and inhibits cell growth. We examined in rats the activity of tissue phosphatases after arterial wall injury and treatment with octreotide and its effect on AP-1 and NF-kappaB phosphorylation and arterial response to injury. The activity of PTP did not change after balloon injury. Treatment of rats with PTP stimulator octreotide increased the PTP activity by 20% +/- 18% in uninjured arteries (p = 0.04 compared with control) and by 49% +/- 44% compared with injured untreated rats (p = 0.017). Treatment of rats with okadaic acid, a specific phosphatase inhibitor, prevented the octreotide-induced increase in PTP activity. PP2A activity of uninjured arteries was not affected significantly with treatment with octreotide (105% +/- 21%, p = 0.57 compared with control). After balloon injury PP2A activity was significantly reduced, 54% +/- 24% of control (p = 0.001). This reduction was prevented with treatment with octreotide, activity 88% +/- 25% of control. When rats were treated with octreotide and okadaic acid, the activity of PP2A in uninjured arteries was decreased to 65% +/- 12% of control (p = 0.03) and the injury-induced reduction was preserved, activity 54% +/- 8% of control (p = 0.001). There was no change in PP2B and C activity after balloon injury. Increased phosphatase activity with octreotide was associated with stabilization of the unphosphorylated form and reduction in nuclear binding of AP-1 and NF-kappaB and was associated with reduced SMC proliferation after balloon injury. Inhibition of increased phosphatase activity with okadaic acid was associated with increased nuclear binding of AP-1 and NF-kappaB. Increased nuclear binding of AP-1 and NF-kappaB after injury was associated with increased expression of fos, jun, and p105 subunit mRNA and restored the proliferative response of SMC after balloon injury. We conclude that the activity of PP2A is decreased after arterial balloon injury which leads to increased AP-1 and NF-kappaB phosphorylation and nuclear binding and is involved in regulation of SMC proliferation. Treatment with octreotide prevented the injury-induced reduction in PP2A activity and decreased transcription factor phosphorylation and SMC proliferation. Modification of phosphatase activity is a potential regulatory mechanism of arterial wall response to injury.

Animals↗
PubMed1998

Studies of the regulation of the mouse carboxyl ester lipase gene in mammary gland.

The lactating mammary gland and pancreas of mouse constitute the main tissues for synthesis and secretion of a bile-salt-stimulated lipase called carboxyl ester lipase (CEL). In this paper we have analysed the endogenous CEL gene expression in mammary gland. It is shown that the gene is expressed at day 14 of pregnancy, which is synchronous with that of the whey acidic protein (WAP) gene. Even though the CEL and WAP genes are induced at the same time during mammary gland differentiation, their regulation is different with respect to dependence on lactogenic hormones. The high induction of the WAP gene expression due to the activation of signal transducer and activator of transcription (STAT)5 by prolactin has not been observed for the CEL gene, even though it has been demonstrated that both STAT5 isoforms interact with one of the gamma-interferon activation sequence sites in the promoter of the CEL gene. Hence we have demonstrated that the prolactin/STAT5 signal is not involved in a general and significant activation of 'milk genes'. Instead of a direct effect of the lactogenic hormones, the up-regulation of the CEL gene is correlated with an increase in the number of differentiated epithelial cells. Furthermore, promoter studies using the mammary-gland-derived cell line, HC11, show that a major positive element in the CEL gene promoter interacts with a member(s) of the CCAAT-binding transcription factor/nuclear factor 1 family, binding to a palindromic site. Binding of this factor(s) is important for the tissue-specific activation of the CEL gene in the mammary gland, because no activation by this factor(s) was seen in cells of pancreatic origin.

Animals↗
PubMed1998

The specificity of the CRM1-Rev nuclear export signal interaction is mediated by RanGTP.

Nuclear export of intron-containing human immunodeficiency virus type 1 (HIV-1) RNA is mediated by the viral Rev protein that contains both an RNA binding domain specific for the viral Rev response element (RRE) and a nuclear export signal (NES). The cellular CRM1 (Exportin1) protein functions as a nuclear export receptor for proteins carrying a Rev-like NES in a process that also requires the GTP bound form of the Ran GTPase. Using purified recombinant factors, we show by co-precipitation, gel mobility shift and protein footprinting assays that full-length Rev protein interacts directly with CRM1 in vitro independently of both the integrity of the characteristic leucine residues of the NES and the presence of the cytotoxin leptomycin B (LMB). Addition of RanGTP induces the formation of an RRE-Rev-CRM1-RanGTP complex that is sensitive to LMB, NES mutations, and Ran being charged with GTP. Within this complex, CRM1 is readily cross-linked to Cys89 near the NES of Rev. By protein footprinting, we demonstrate that the NES of Rev and two regions in CRM1 become inaccessible to endoproteinases upon binding suggesting that these regions are involved in protein-protein interactions. Our data are consistent with a model in which CRM1 is the nuclear export receptor for the Rev-RRE ribonucleoprotein complex and that RanGTP binds to a preformed Rev-CRM1 complex and specifies a functional interaction with the NES.

Base Sequence↗
PubMed1998

Transcriptional regulation of the human carboxyl ester lipase gene in exocrine pancreas. Evidence for a unique tissue-specific enhancer.

The human carboxyl ester lipase (CEL) is an important enzyme for the intestinal absorption of dietary lipids. The gene is highly expressed in exocrine pancreas and in the mammary gland during pregnancy and lactation. In this paper, we have focused on its transcriptional regulation in exocrine pancreas. Reporter gene analysis in cell cultures reveals that a high level of tissue-specific expression is established by the proximal 839 base pairs of the 5'-flanking region. This is due to a strong enhancer, located at -672 to -637. Transfections in mammary gland-derived cells reveal that the enhancer is pancreas-specific and does not contribute to the mammary gland expression. This indicates that the expression of the CEL gene in the mammary gland and pancreas, respectively, is due to two different regulatory systems. Further characterizations of the enhancer reveal that it is composed of two closely located cis-elements. The proximal element mediates a positive effect, whereas the distal element exerts a silencing effect on the positive proximal element. The functional enhancer complex is composed of ubiquitously expressed factors, since similar interactions are achieved with nuclear extracts from cells derived from other tissues. However, no enhancer activity is achieved in such cells. Hence, the net enhancer activity is the result of a tissue-specific balance between factors interacting with the two elements. Since none of the described cis-elements show any clear homology to known cis-elements, we propose that the interacting complex is composed of yet unidentified transcription factors.

Base Sequence↗
PubMed1998

Treatment effects on serum lipoprotein lipids, apolipoproteins and low density lipoprotein particle size and relationships of lipoprotein variables to progression of coronary artery disease in the Bezafibrate Coronary Atherosclerosis Intervention Trial (BECAIT).

OBJECTIVES: To investigate the mechanisms by which bezafibrate retarded the progression of coronary lesions in the Bezafibrate Coronary Atherosclerosis Intervention Trial (BECAIT), we examined the relationships of on-trial lipoproteins and lipoprotein subfractions to the angiographic outcome measurements. BACKGROUND: BECAIT, the first double-blind, placebo-controlled, randomized serial angiographic trial of a fibrate compound, showed that progression of focal coronary atherosclerosis in young survivors of myocardial infarction could be retarded by bezafibrate treatment. METHODS: A total of 92 dyslipoproteinemic men who had survived a first myocardial infarction before the age of 45 years were randomly assigned to treatment for 5 years with bezafibrate (200 mg three times daily) or placebo; 81 patients underwent baseline and at least one post-treatment coronary angiography. RESULTS: In addition to the decrease in very low density lipoprotein (VLDL) cholesterol (-53%) and triglyceride (-46%) and plasma apolipoprotein (apo) B (-9%) levels, bezafibrate treatment resulted in a significant increase in high density lipoprotein-3 (HDL3) cholesterol (+9%) level and a shift in the low density lipoprotein (LDL) subclass distribution toward larger particle species (peak particle diameter +032 nm). The on-trial HDL3 cholesterol and plasma apo B concentrations were found to be independent predictors of the changes in mean minimum lumen diameter (r=-0.23, p < 0.05), and percent (%) stenosis (r = 0.30, p < 0.01), respectively. Decreases in small dense LDL and/or VLDL lipid concentrations were unrelated to disease progression. CONCLUSIONS: Our results suggest that the effect of bezafibrate on progression of focal coronary atherosclerosis could be at least partly attributed to a rise in HDL3 cholesterol and a decrease in the total number of apo B-containing lipoproteins.

Adult↗
PubMed1998

Picoliter sample preparation in MALDI-TOF MS using a micromachined silicon flow-through dispenser.

This paper presents a picoliter sample preparation technique utilizing the flow-through principle, allowing on-line coupling of chromatographic systems to be made. The work was performed in order to investigate the characteristics and the physicochemical properties of the sample preparation using typical mobile phase conditions from mu-CLC (column liquid chromatography) separations. The device presented here is a pressure pulse-driven dispenser, formed by two silicon structures processed by conventional micromachining. The pressure pulse is generated in the flow-through channel by a piezoceramic element. Depending on the orifice size, the droplets ejected range between 30 and 200 pL. The maximum ejection frequency is 500 Hz, limited by resonances within the unit. A pyramid-shaped nozzle improves the directivity of the droplets since it reduces the wetting of the orifice front surface area. The risk of particles sticking close to the orifice is also minimized. The analyses of the deposited sample spots were carried out on a matrix-assisted laser desorption/ionization time-of-flight mass spectrometer with delayed extraction. It was possible to detect attomole amounts (159-248 amol) of various proteins (cytochrome c, ribonuclease A, lysozyme, and myoglobin) from a single droplet of matrix:analyte 1:1 (drop volume approximately 110 pL). Additionally, it was found that sample enrichment could be carried out using multiple depositions on the same spot; i.e., 31 nM of insulin was easily detected when more than four depositions were made on the same spot, while no detection was possible without sample enrichment. Size optimization of the MALDI sample spot gave target zones of 100-500-micron diameter that matched the size of the laser focal point and resulted in a considerably increased sample throughput.

Cell Line↗
PubMed1998

Mechanisms of bupivacaine action on Na+ and K+ channels in myelinated axons of Xenopus laevis.

The local anaesthetic bupivacaine has recently been proposed to inhibit Na+ channels indirectly by making the resting potential less negative. To test this hypothesis we analysed the effects of bupivacaine on voltage and current clamped nodes of Ranvier. Contrary to the hypothesis, the leak current and the resting potential were unaffected. The Na+ and K+ channels were, however, affected at relatively low concentrations (33 microM). Steady-state activation curves were decreased without notable shift effects, whereas the Na+ inactivation curve was decreased and shifted in negative direction. The effect on the Na+ current was tentatively explained by a single-site, state-dependent binding model (Kd = 44 microM), while that on the K+ current was explained by two population-specific mechanisms, one open-state dependent (Kd = 550 microM) and one state independent (Kd = 59 microM). The binding stoichiometry was higher than 1:1 for the main sites of action. In conclusion, bupivacaine exerts its main anaesthetic action on myelinated nerve axons by a direct modification of Na+ channels.

Anesthetics, Local↗
PubMed1998

Sample enrichment in a single levitated droplet for capillary electrophoresis.

This paper describes sample enrichment in a single levitated droplet for capillary electrophoresis (CE) analysis. The droplet was trapped in an acoustical field. The minute sample volumes needed for the enrichment procedure were precisely handled using a piezoelectric flow-through liquid microdispenser. Droplets with a volume of 65 pl were ejected from the device at a repetition rate ranging from one single droplet up to several hundreds per second. By counting the number of droplets ejected and accumulated in the levitated drop the sample volume was controlled. Through solvent evaporation the analytes were enriched in the diminishing droplet. The droplet was then injected into a CE capillary and the analytes, dansyl-Gly and dansyl-Val dissolved in ethanol, were separated in a 100 mM borate buffer (pH 9.0) utilising UV-absorption detection at 200 nm near the capillary outlet. Enrichment of 36000 sample droplets (2.3 microl) through solvent evaporation in the levitated drop resulted in a concentration limit of detection (CLOD) of 15 nM for the dansylated amino acids as compared to a CLOD of 2.5 microM which was achieved using standard hydrodynamic injection without preconcentration.

Electrophoresis, Capillary↗