| 1. |
- Cain, Peter A, et al.
(författare)
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Age and gender specific normal values of left ventricular mass, volume and function for gradient echo magnetic resonance imaging: a cross sectional study.
- 2009
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Ingår i: BMC medical imaging. - : Springer Science and Business Media LLC. - 1471-2342. ; 9:2
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Tidskriftsartikel (refereegranskat)abstract
- Knowledge about age-specific normal values for left ventricular mass (LVM), end-diastolic volume (EDV), end-systolic volume (ESV), stroke volume (SV) and ejection fraction (EF) by cardiac magnetic resonance imaging (CMR) is of importance to differentiate between health and disease and to assess the severity of disease. The aims of the study were to determine age and gender specific normal reference values and to explore the normal physiological variation of these parameters from adolescence to late adulthood, in a cross sectional study.
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| 2. |
- Cain, Peter, et al.
(författare)
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Physiological determinants of the variation in left ventricular mass from early adolescence to late adulthood in healthy subjects
- 2005
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Ingår i: Clin Physiol Funct Imaging. - 1475-0961. ; 25:6, s. 332-9
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: The physiological determinants of left ventricular mass (LVM) measured by cardiac magnetic resonance (CMR) imaging are not well defined as prior investigators have studied either adults or adolescents in isolation or have not strictly excluded hypertension or accounted for the effects of exercise habits, haemodynamic, demographic, or body shape characteristics. METHODS: A total of 102 healthy volunteers (12-81 years, 53 males) underwent CMR. All parameters [unstandardized and adjusted for body surface area (BSA)] were analysed according to gender and by adolescence versus adulthood (adolescents <20 years, adults > or = 20 years). The influence of haemodynamic factors, exercise, and demographic factors on LVM were determined with multivariate linear regression. Results: LVM rose during adolescence and declined in adulthood. LVM and LVMBSA were higher in males both in adults (LVM: 188 +/- 22 g versus 139 +/- 21 g, P < 0.001; LVMBSA: 94 +/- 11 g m(-2) versus 80 +/- 11 g m(-2), P < 0.001) and in adolescents when adjusted for BSA (LVM: 128 +/- 29 g versus 107 +/- 20 g, P = 0.063; LVMBSA: 82 +/- 8 g m(-2) versus 71 +/- 10 g m(-2), P = 0.025). In adults, systolic blood pressure (SBP) and self-reported physical activity increased while meridional and circumferential wall stress were constant with age. Multivariate regression analysis revealed age, gender, and BSA as the major determinants of LVM (global R2 = 0.69). CONCLUSIONS: Normal LVM shows variation over a broad age range in both genders with a rise in adolescence and subsequent decline with increasing age in adulthood despite an increase in SBP and physical activity. BSA, age, and gender were found to be major contributors to the variation in LVM in healthy adults, while haemodynamic factors, exercise, and wall stress were not.
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| 3. |
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| 4. |
- Dainiak, Maria, et al.
(författare)
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Improved methods for prepurification and detection of Staphylococcal Enterotoxin B from cell-free culture filtrate
- 2005
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Ingår i: Biotechnology Progress. - : Wiley. - 1520-6033 .- 8756-7938. ; 21:4, s. 1347-1351
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Tidskriftsartikel (refereegranskat)abstract
- An improved ELISA method for the detection of Staphylococcal. Enterotoxin B (SEB) in protein A preparations is presented. Fab fragments were obtained by digestion with papain of anti-SEB IgG bound to SEB immobilized on Sepharose 4B. Anti-SEB and peroxidase-labeled Fab fragments from secondary antibodies were successfully used in a modified ELISA of SEB in protein A preparations. SEB-Sepharose was used repeatedly for the production of anti-SEB Fab fragments by papain digestion without loss of affinity. In addition, for the purification of SEB from crude culture filtrates, an initial step utilizing a combined heat and pH treatment for the removal of significant amounts of contaminating proteins without losses of toxin activity is presented. This pretreatment step yielded positive effects in further downstream processing considering both shortened time and an increase in total recovery.
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| 5. |
- Deraz, Sahar, et al.
(författare)
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Production and physicochemical characterization of acidocin D20079, a bacteriocin produced by Lactobacillus acidophilus DSM 20079
- 2007
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Ingår i: World Journal of Microbiology & Biotechnology. - : Springer Science and Business Media LLC. - 0959-3993 .- 1573-0972. ; 23:7, s. 911-921
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Tidskriftsartikel (refereegranskat)abstract
- Lactobacillus acidophilus DSM 20079 is the producer of a novel bacteriocin termed acidocin D20079. In this paper, a partial sequence of this peptide is determined, together with data on its secondary structure. A modification of the MRS-growth medium (replacing the detergent Tween 80 with oleic acid), was shown to improve the production level of the peptide by one order of magnitude, as well as to stabilize the activity level. Addition of a detergent (Tween 20, less interfering in mass spectrometric analysis), was however necessary for solubilization of the purified acidocin D20079. Digestion of the peptide followed by de-novo sequencing of generated fragments, allowed determination of a partial sequence consisting of 39 of the totally estimated 65 residues. Acidocin D20079 has a high content of glycine residues, hydrophobic residues, and acidic residues. No modified amino acids were found. Edman degradation, and C-terminal sequencing failed, suggesting that the peptide may be cyclic, and a novel member of class IIc bacteriocins. Circular dichroism spectroscopy and secondary structure prediction showed random coil conformation in aqueous solution, but secondary structure was induced in the presence of sodium-dodecyl sulfate. The data could be fitted assuming 2-13% of the residues to be in alpha-helix and 23-27% of the residues to be in beta-strand conformation. This indicates that a membrane/membrane-mimicking hydrocarbon-water interface induces an active conformation.
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| 6. |
- Deraz, Sahar, et al.
(författare)
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Purification and characterisation of acidocin D20079, a bacteriocin produced by Lactobacillus acidophilus DSM 20079
- 2005
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Ingår i: Journal of Biotechnology. - : Elsevier BV. - 1873-4863 .- 0168-1656. ; 117:4, s. 343-354
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Tidskriftsartikel (refereegranskat)abstract
- Bacteriocins are natural antimicrobial agents produced by food fermentative bacteria. Lactobacillus acidophilus DSM 20079 produces a small bacteriocin, with a molecular mass of 6.6 kDa, designated acidocin D20079. This antimicrobial peptide was extremely heat-stable (30 min at 121 degrees C) and was active over a wide pH range. It was found to be sensitive to proteolytic enzymes (trypsin, ficin, pepsin, papain, and proteinase K). Acidocin D20079 has a narrow inhibitory spectrum restricted to the genus Lactobacillus which includes L. sakei NCDO 2714, an organism known to cause anaerobic spoilage of vacuum-packaged meat. Maximum production of acidocin D20079 in MRS broth was detected at pH 6.0, and the peptide was purified by ammonium sulphate precipitation followed by sequential cation exchange and hydrophobic interaction chromatography. Purified acidocin D20079 spontaneously formed spherulite crystals during dialysis. As the N-terminus was found to be blocked for sequencing, matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry was used to determine a partial sequence. and the molecular mass of the bacteriocin in the formed crystals (6.6 kDa). Estimates of the molecular weight of the partially purified peptide, using tricine-SDS-PAGE, in which bacteriocin activity was confirmed by overlayer techniques were in accordance with this value. (c) 2005 Published by Elsevier B.V.
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| 7. |
- Elovson Grey, Carl, et al.
(författare)
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A mass spectrometric investigation of native and oxidatively inactivated chloroperoxidase
- 2007
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Ingår i: ChemBioChem. - : Wiley. - 1439-4227 .- 1439-7633. ; 8:9, s. 1055-1062
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Tidskriftsartikel (refereegranskat)abstract
- The enzyme chloroperoxidase (CPO) found in Calclariomyces fumago is able to catalyze several stereoselective oxidation reaction by using a dean oxidant, usually hydrogen peroxide (H2O2), without the need for expensive cofactor generation. CPO's lack of operational stability however, is a major limitation for its commercial use. In the present study, a capillary-LC on-line trypsin- digestion system combined with reversed-phase chromatography and mass spectrometric detection was optimized for studying the primary sequence of CPO. Samples containing native CPO, CPO treated with H2O2, and CPO oxidatively inactivated by the use of indole and H2O2 were analyzed and compared. Three oxidized peptides were found in the samples treated with H2O2. Two additional oxidized peptides were found in the CPO samples that were completely inactivated, one of which contained an oxidized cysteine residue, Cys50, which is an essential amio acid due to its function as the axial ligand to the iron in the heme - the prosthetic group in CPO. In addition, the heme group was absent in the inactivated samples but was readily detected in other samples.
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| 8. |
- Engblom, Henrik, et al.
(författare)
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The endocardial extent of reperfused first-time myocardial infarction is more predictive of pathologic Q waves than is infarct transmurality: a magnetic resonance imaging study.
- 2007
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Ingår i: Clinical Physiology and Functional Imaging. - 1475-0961. ; 27:2, s. 101-108
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Tidskriftsartikel (refereegranskat)abstract
- Historically, Q-wave myocardial infarction (MI) has been equated with transmural MI. This association have, however, recently been rejected. The endocardial extent of MI is another potential determinant of pathological Q waves, since the first part of the QRS complex where the Q wave appears reflects depolarization of subendocardial myocardium. Therefore, the aim of the present study was to test the hypothesis that endocardial extent of MI is more predictive of pathological Q waves than is MI transmurality and to investigate the relationship between QRS scoring of the ECG and MI characteristics. Twenty-nine patients with reperfused first-time MI were prospectively enrolled. One week after admission, delayed contrast-enhanced magnetic resonance imaging (DE-MRI) was performed and 12-lead ECG was recorded. Size, transmurality and endocardial extent of MI were assessed by DE-MRI. Q waves were identified with Minnesota coding and electrocardiographic MI size was estimated by QRS scoring of the ECG. There was a significant difference between patients with and without Q waves with regard to MI size (P = 0.03) and endocardial extent of MI (P = 0.01), but not to mean and maximum MI transmurality (P = 0.09 and P = 0.14). Endocardial extent was the only independent predictor of pathological Q waves. Endocardial extent of MI was most strongly correlated to QRS score (r = 0.86, P < 0.001) of the MI variables tested. The endocardial extent of reperfused first-time acute MI is more predictive of pathological Q waves than is MI transmurality.
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| 9. |
- Hanora, Amro, et al.
(författare)
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Capture of bacterial endotoxins using a supermacroporous monolithic matrix with immobilized polyethyleneimine, lysozyme or polymyxin B
- 2005
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Ingår i: Journal of Biotechnology. - : Elsevier BV. - 1873-4863 .- 0168-1656. ; 118:4, s. 421-433
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Tidskriftsartikel (refereegranskat)abstract
- Bacterial endotoxins (BEs) are integrated part of Escherichia coli, a microorganism widely used for the production of recombinant proteins. BEs should be eliminated in the course of down stream processing of target protein produced by these bacteria. Supermacroporous monolith (continuous bed) columns, so called cryogel columns, with immobilized polyethyleneimine (PEI), polymyxin B (PMB) and lysozyme were employed for BEs capture. Due to the large interconnected pores it was possible to use cryogel columns at flow rates as high as 10 ml/min. The columns packed with Sepharose CL-4B with immobilized PEI, PMB and lysozyme were impossible to use at these high flow rates due to the collapse of the bed. The dynamic capacities of the cryogel columns were nearly independent of the flow rate. In the presence of EDTA, BEs were quantitatively captured from mixtures with a model protein, bovine serum albumin (BSA) at pH 7.2 with practically no protein losses. At pH 3.6 BEs were captured directly from non-clarified E. coli cell lysate resulting in more than 104 times BEs clearance. (C) 2005 Elsevier B.V. All rights reserved.
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| 10. |
- Hedström, Gustaf, et al.
(författare)
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Mast cell infiltration is a favourable prognostic factor in diffuse large B-cell lymphoma
- 2007
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Ingår i: British Journal of Haematology. - : Wiley. - 0007-1048 .- 1365-2141. ; 138:1, s. 68-71
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Tidskriftsartikel (refereegranskat)abstract
- Previous studies indicate that the inflammatory response in diffuse large B-cell lymphomas (DLBCL) is important for the clinical outcome. Mast cells are key regulators in this response; we investigated whether the number of tryptase-positive mast cells is correlated with clinical outcome. Patients with many mast cells had a significantly better event-free survival (EFS) compared to those with few mast cells (P < 0.03 in both germinal centre (GC) and non-GC DLBCL. This supports the idea that the infiltration of mast cells is a reflection of the host inflammatory response and is related to a favourable outcome.
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| 11. |
- Hedström, Martin, et al.
(författare)
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Continuous measurements of a binding reaction using a capacitive biosensor
- 2005
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Ingår i: Biosensors & Bioelectronics. - : Elsevier BV. - 1873-4235 .- 0956-5663. ; 21:1, s. 41-48
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Tidskriftsartikel (refereegranskat)abstract
- A capacitive biosensor with polyclonal antibodies raised against human serum albumin (HSA) immobilized on a gold transducer has been developed for continuous measurement of HSA in the μM-range. A mathematical model has been refined to describe integral HSA-binding curves assuming that (i) binding is essentially irreversible under the conditions used, (ii) the signal is scaled as the number of non-occupied binding sites and (iii) the rate of disappearance of available binding sites is scaled as the number of available binding sites and analyte concentration in solution. Deconvolution of the curves using the mathematical model indicates clearly that it is possible to retrieve concentration profiles (isocratic, linearly or exponentially increasing gradients) of the analyte in the continuous sample flow from the normalized integral binding (NIB) curves. The data presented constitutes the theoretical background and the first step towards the development of an analytical system allowing on-line detection of the concentration profile of the analyte from NIB-curves. Since the system can be used for extended time periods between regeneration steps, a low frequency of regeneration steps can be expected.
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| 12. |
- Hedström, Martin, et al.
(författare)
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Fast on-column protein digestion with subsequent peptide mapping using tandem mass spectrometry with information dependent acquisition
- 2005
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Ingår i: Journal of Chromatography A. - : Elsevier BV. - 0021-9673. ; 1080:2, s. 117-123
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Tidskriftsartikel (refereegranskat)abstract
- A platform for rapid on-line protein digestion of protein mixtures for direct infusion to a mass spectrometer is presented. A mixture of protein A, staphylococcal enterotoxin B and cytochrome c was used as a model mixture injected on a gel filtration column and a trypsin reactor which were connected in series to a micro liquid chromatography (mu LC) system. The peptides in the column eluate were analyzed with ESI tandem mass spectrometry, utilizing information dependent acquisition (IDA). In one step, the proteins in the mixture (mu M concentrations) were concomitantly desalted, separated, digested and identified with an overall analysis time of less than 40 min. Protein sequence coverage of 78-95% for the involved substances was achieved. (c) 2005 Elsevier B.V. All rights reserved.
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| 13. |
- Hedström, Martin, et al.
(författare)
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Miniaturized on-line digestion system for the sequential identification and characterization of protein analytes
- 2007
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Ingår i: Journal of Chromatography A. - : Elsevier BV. - 0021-9673. ; 1146:1, s. 17-22
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Tidskriftsartikel (refereegranskat)abstract
- A miniaturized on-column digestion system constructed for the sequential analysis of semi-purified protein analytes is presented. By utilizing fused silica capillary (diameter 150 mu m) packed with a zone of trypsin-modified Eupergit C beads and a second zone of reversed-phase C18 material, a linear column set-up was constructed. The protein analytes (pmol amounts) were first digested in the 600 nl trypsin reactor portion of the system. Next, the generated peptides were trapped in the C18 column shaped as an electrospray emitter. Finally, after washing the matrix free from salts and other hydrophilic impurities present in the sample, peptides were eluted. A stepwise increased concentration profile of organic solvent, created by a dual syringe pump system, promoted the release of bound peptides, which were identified by electrospray ionization MS/MS. This approach proved to be very efficient, achieving almost complete digestion of the proteins studied, with suitable operational stability maintained for more than 1 week. Further, a small nebulizer was designed and fitted to the electrospray emitter. A significant improvement of the spray stability was observed and droplet build-up on the capillary was avoided, even at flow rates well above 1500 nl/min. The proteins chloroperoxidase, staphylococcal enterotoxin B and protein A (injection volume 0.3 mu l, salt concentration 0.2-1 M) were sequentially digested, desalted, eluted, detected and conclusively identified by bioinformatics web tools with an analytical cycle time of 10 min.
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| 14. |
- Hedström, Martin, et al.
(författare)
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Monolithic macroporous albumin/chitosan cryogel structure: a new matrix for enzyme immobilization.
- 2008
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Ingår i: Analytical and Bioanalytical Chemistry. - : Springer Science and Business Media LLC. - 1618-2642 .- 1618-2650. ; 390:3, s. 907-912
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Tidskriftsartikel (refereegranskat)abstract
- A novel monolithic macroporous material was developed by cross-linking hen egg albumin (HEA) and chitosan with glutaraldehyde at subzero temperatures. A macroporous cryogel structure allowed efficient mass transport of solutes within the material. In one application, albumin was partially replaced with active enzymes (glucose oxidase and horseradish peroxidase) resulting in the production of macroporous biocatalyst preparations suitable for flow-injection analysis of glucose in the low millimolar range. In another application, the proteolytic enzymes savinase and esperase were coupled to the macroporous structure via free amino groups on the pore walls using glutaraldehyde as cross-linker/spacer agent. The low hydraulic resistance of the matrix allowed for the development of a generic, high-performance online protein digestion system utilizing the wall-bound proteases.
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| 15. |
- Hedström, Martin
(författare)
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Process Analytical Tools for the Monitoring and Characterization of Bacterial Toxins
- 2006
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- In the era of proteomics, glycomics and ultimately metabolomics, with constantly increasing knowledge regarding the human biosystem, the number of specialized therapeutic agents which will be developed against different diseases is predicted to be on the upswing. An increasing portion of these new drugs will, according to recent predictions (Labrou, 2003), be of proteinaceous origin produced according to biotechnological standards. Hence, down stream processes (DSP) in which large amounts of specific target proteins are to be processed and purified, with high demands on process monitoring and control, will be of further increasing importance in the future. In the light of these predictions, the U.S. Food and Drug Administration (FDA) has proposed a set of new initiatives that primarily address the development of process analytical technology (PAT) as a concept within pharmaceutical manufacturing. The idea with PAT is to stimulate creative strategies for improved process developments, which in the end should lead to increased productivity concomitantly with the assurance of acceptable end-product quality (U.S. FDA, 2002). The definition of PAT involves the use of an overall process optimisation strategy applied at all production levels. Hence, the utilization of advanced bioprocess monitoring tools, which could lead to reduced production cycle times and prevention of rejects, scrap, and the need for re-processing, is covered in the description. A recent tendency within bioproduction monitoring is the shift in focus from the target molecule itself towards an increased control of the impurities present (World Health Organization, 1999). Among the relevant contaminants typically present in a bioproduction process, bacterial toxins have received high priority from the regulatory agencies (Ecker et al., 2005). In fact, a clear analytical strategy for the control of e.g. bacterial endotoxins is a necessity for the recombinant production of any FDA-approved pharmaceutically active proteinaceous agent (U.S. FDA, 1985). Currently, the only available method for endotoxin detection is the limulus amebocyte lysate (LAL) assay which is based on the Limulus lectin derived from the American horseshoe crab,. The LAL assay is expensive and time-consuming. Moreover, it is non-functional for the detection of endotoxins in trace amounts. Hence, the development of new, less labour-intensive methods with high sensitivity and specificity for the direct detection of endotoxins is highly desirable. Another important category of contaminants associated with biotechnological production comprises the exotoxins. This class of bacterial toxins typically includes soluble proteinaceous compounds, continually released by living bacteria. Extreme toxicities together with a typical ability to retain high activity under harsh conditions and in dilute solutions make exotoxins exceptionally competent as poisons, often with high mortality rates. The treatment of toxic by-products in biotechnological production is strictly regulated and the strategies for the removal of toxins associated with the product are often complex and completely dependant on the product application. A parallel situation is the handling of waste liquids remaining from the process. Fermentation of cell strains, capable of excreting exotoxic compounds to the surroundings, will consequently give waste liquids that require detoxification before being discarded. Hence, cost-effective methods for performing such tasks could be of interest to the manufacturer as e.g. the cost of sending the material for incineration could be a heavy toll on the profit margins. The objective in this thesis work was to investigate different means to analyze, characterize and process selected toxins of bacterial origin related to biotechnological production. With a vast array of compounds falling into the category of bacterial toxins, two different types were chosen; staphylococcal enterotoxin B (SEB), a 23 kDa exotoxin produced by Staphylococcus aureus and the heterogeneous group of endotoxins derived from the outer cell-wall of the gram-negative bacteria Escherichia coli.
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| 16. |
- Heiberg, Einar, et al.
(författare)
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Semi-automatic quantification of myocardial infarction from delayed contrast enhanced magnetic resonance imaging
- 2005
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Ingår i: Scandinavian Cardiovascular Journal. - : Informa UK Limited. - 1401-7431 .- 1651-2006. ; 39:5, s. 267-275
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Tidskriftsartikel (refereegranskat)abstract
- Objective. Accurate and reproducible assessment of myocardial infarction is important for treatment planning in patients with ischemic heart disease. This study describes a novel method to quantify myocardial infarction by semi-automatic delineation of hyperenhanced myocardium in delayed contrast enhanced (DE) magnetic resonance (MR) images. Design. The proposed method automatically detects the hyperenhanced tissue by first determining the signal intensity of non-enhanced myocardium. A fast level set algorithm was used to limit the heterogeneity of the hyperenhanced regions, and to exclude small regions that constitute noise rather than infarction. The method was evaluated in 40 patients, 20 with acute infarction and 20 with chronic healed infarction using scanners from two different manufacturers. Infarct size measured by the proposed semi-automatic method was compared with manual measurements from three experienced observers. The software used is freely available for research purposes at http://segment.heiberg.se. Results. The difference in infarct size between semi-automatic quantification and the mean of the three observers was 6.1 ± 6.6 ml (mean ± SD), and the interobserver variability (SD) was 4.2 ml. Conclusions. The method presented is a highly automated method for analyzing myocardial viability from DE-MR images. The bias of the method is acceptable and the variability is in the same order of magnitude as the interobserver variability for manual delineations. © 2005 Taylor & Francis.
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| 17. |
- Labib, Mahmoud, et al.
(författare)
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A capacitive biosensor for detection of staphylococcal enterotoxin B.
- 2009
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Ingår i: Analytical and Bioanalytical Chemistry. - : Springer Science and Business Media LLC. - 1618-2642 .- 1618-2650. ; 393, s. 1539-1544
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Tidskriftsartikel (refereegranskat)abstract
- A sensitive method for the detection of staphylococcal enterotoxin B (SEB) using a flow-injection capacitive biosensor is presented. SEB was purified from a crude culture filtrate of Staphylococcus aureus through three chromatographic steps. The first two steps were based on ion-exchange chromatography, and the last step was carried out on a gel filtration column. The SEB recovery values after the purification stages were 88%, 74%, and 12%, respectively. A horseradish peroxidase labeled antistaphylococcal enterotoxin B was prepared by the periodate method and was further employed in a sandwich-enzyme-linked immunosorbent assay (ELISA) for the determination of SEB concentrations in different samples obtained during the processing of the crude filtrate. The capacitive biosensor could detect SEB concentrations as low as 0.3 pg ml(-1) with a linearity ranging from 2.8 pg ml(-1) to 2.8 ng ml(-1) under optimized conditions. The response time was about 10 min. A good agreement was achieved between the developed capacitive biosensor system and ELISA as a reference method for detection of SEB levels in different purification samples. The newly developed sensor has the benefits of simplicity, high sensitivity, and multiple use capability.
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| 18. |
- Labib, Mahmoud, et al.
(författare)
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A capacitive immunosensor for detection of cholera toxin
- 2009
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Ingår i: Analytica Chimica Acta. - : Elsevier BV. - 1873-4324 .- 0003-2670. ; 634:2, s. 255-261
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Tidskriftsartikel (refereegranskat)abstract
- Contamination of food with biological toxins aswell as their potential use asweapons of mass destruction has created an urge for rapid and cost effective analytical techniques capable of detecting trace amounts of these toxins. This paper describes the development of a sensitive method for detection of cholera toxin (CT) using a flow-injection capacitive immunosensor based on self-assembled monolayers. The sensing surface consists of monoclonal antibodies against the B subunit of CT (anti-CT), immobilized on a gold transducer. Experimental results showthat the immunosensor responded linearly to CT concentrations in the range from 1.0×10−13 to 1.0×10−10 M under optimized conditions. The limit of detection (LOD) was 1.0×10−14 M. Two more analytical methods were employed for detection of CT using the same antibody namely, sandwich ELISA and surface plasmon resonance (SPR)-based immunosensor. The former had an LOD of 1.2×10−12 M and a working range from3.7×10−11 to 2.9×10−10 M whereas, the latter had an LOD of 1.0×10−11 M and a linearity ranging from 1.0×10−9 to 1.0×10−6 M. These results demonstrate that the developed capacitive immunosensor system has a higher sensitivity than the other two techniques. The binding affinity of CT to the immobilized anti-CT was determined using the SPR-based immunosensor and an association constant (KA) of 1.4×109 M−1 was estimated.
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| 19. |
- Labib, Mahmoud, et al.
(författare)
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A multipurpose capacitive biosensor for assay and quality control of human immunoglobulin G.
- 2009
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Ingår i: Biotechnology and Bioengineering. - : Wiley. - 1097-0290 .- 0006-3592. ; 104, s. 312-320
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Tidskriftsartikel (refereegranskat)abstract
- We report a flow-injection biosensor system with a capacitive transducer for assay and quality control of human immunoglobulin G (hIgG). The sensing platform is based on self-assembled monolayers (SAMs) of carboxylic acid terminated alkyl-thiols with covalently attached concanavalin A. The electrochemical characteristics of the sensor surface were assessed by cyclic voltammetry using a permeable redox couple (potassium ferricyanide). The developed biosensor proved capable of performing a sensitive label-free assay of hIgG with a detection limit of 1.0 microg mL(-1). The capacitance response depended linearly on hIgG concentration over the range from 5.0 to 100 microg mL(-1), in a logarithmic plot. Typical measurements were performed in 15 min and up to 18 successive assays were achieved without significant loss of sensitivity using a single electrode. In addition, the biosensor can detect hIgG aggregates with concentrations as low as 0.01% of the total hIgG content (5.0 microg mL(-1)). Hence, it represents a potential post-size-exclusion chromatography-UV (post-SEC-UV) binding assay for in-process quality control of hIgG, which cannot be detected by SEC-UV singly at concentrations below 0.3% of the total hIgG content. Biotechnol. Bioeng. (c) 2009 Wiley Periodicals, Inc.
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| 20. |
- Limbut, Warakorn, et al.
(författare)
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Capacitive biosensor for detection of endotoxin
- 2007
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Ingår i: Analytical and Bioanalytical Chemistry. - : Springer Science and Business Media LLC. - 1618-2642 .- 1618-2650. ; 389:2, s. 517-525
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Tidskriftsartikel (refereegranskat)abstract
- A capacitive biosensor for the detection of bacterial endotoxin has been developed. Endotoxin-neutralizing protein derived from American horseshoe crab was immobilized to a self-assembled thiol layer on a biosensor transducer (Au). Upon injection of a sample containing endotoxin, a decrease in the observed capacitive signal was registered. Endotoxin could be determined under optimum conditions with a detection limit of 1.0 x 10(-13) M and linearity ranging from 1.0 x 10(-13) to 1.0 x 10(-10) M. Good agreement was achieved when applying endotoxin preparations purified from an Escherichia coli cultivation to the capacitive biosensor system, utilizing the conventional method for quantitative endotoxin determination, the Limulus amebocyte lysate test as a reference. The capacitive biosensor method was statistically tested with the Wilcoxon signed rank test, which proved the system is acceptable for the quantitative analysis of bacterial endotoxin (P < 0.05).
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| 21. |
- Turner, Pernilla, et al.
(författare)
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Expression, purification, crystallisation and preliminary X-ray diffraction analysis of Thermotoga neapolitana beta-glucosidase B
- 2007
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Ingår i: Acta Crystallographica. Section F: Structural Biology and Crystallization Communications. - 2053-230X. ; 63:9, s. 802-806
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Tidskriftsartikel (refereegranskat)abstract
- -Glucosidases belong to families 1, 3 and 9 of the glycoside hydrolases and act on cello-oligosaccharides. Family 1 and 3 enzymes are retaining and are reported to have transglycosylation activity, which can be used to produce oligosaccharides and glycoconjugates. Family 3 enzymes are less well characterized than their family 1 homologues and to date only two crystal structures have been solved. Here, the expression, purification, crystallization and X-ray diffraction data of a family 3 -glucosidase from the hyperthermophilic bacterium Thermotoga neapolitana are reported. Crystals of selenomethionine-substituted protein have also been grown. The crystals belong to space group C2221, with unit-cell parameters a = 74.9, b = 127.0, c = 175.2 Å. Native data have been collected to 2.4 Å resolution and the structure has been solved to 2.7 Å using the selenomethionine MAD method. Model building and refinement of the structure are under way.
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| 22. |
- Törnvall, Ulrika, et al.
(författare)
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Mass spectrometric analysis of peptides from an immobilized lipase: focus on oxidative modifications.
- 2009
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Ingår i: Rapid Communications in Mass Spectrometry. - : Wiley. - 1097-0231 .- 0951-4198. ; 23:18, s. 2959-2964
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Tidskriftsartikel (refereegranskat)abstract
- Liquid chromatography/tandem mass spectrometry (LC/MS/MS) was used to study the primary structure of immobilized Candida antarctica lipase B (Novozym(R)435) without detaching the enzyme from the carrier. The immobilized enzyme packed in a miniature column was subjected to proteolysis and the peptides released were injected into the mass spectrometer for analysis. The set-up was utilized to determine amino acid oxidation after treatment of the biocatalyst with hydrogen peroxide. In total, sequence coverage of more than 90% was obtained, containing almost all of the amino acids sensitive to oxidation. Oxidation of methionine, tryptophan and cystine residues was observed. The flow system also allowed evaluation of the enzyme activity prior to peptide analysis. The developed method is general and should be applicable to other immobilized enzyme systems and to different treatments.
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