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- Chen, Y. C., et al.
(författare)
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Fluorescence anisotropy studies of molecularly imprinted polymers
- 2006
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Ingår i: Luminescence (Chichester, England Print). - : Wiley. - 1522-7235 .- 1522-7243. ; 21:1, s. 7-14
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Tidskriftsartikel (refereegranskat)abstract
- A molecularly imprinted polymer (MIP) is a biomimetic material that can be used as a biochemical sensing element. We studied the steady-state and time-resolved fluorescence and fluorescence anisotropy of anthracene-imprinted polyurethane. We compared MIPs with imprinted analytes present, MIPs with the imprinted analytes extracted, MIPs with rebound analytes, non-imprinted control polymers (non-MIPs) and non-MIPs bound with analytes to understand MIP’s binding behaviour. MIPs and non-MIPs had similar steady-state fluorescence anisotropy in the range 0.11-0.24. Anthracene rebound in MIPs and non-MIPs had a fluorescence lifetime of tau = 0.64 ns and a rotational correlation time of 0, = 1.2-1.5 ns, both of which were shorter than that of MIPs with imprinted analytes present (tau = 2.03 ns and phi(F) = 2.7 ns). The steady-state anisotropy of polymer solutions increased exponentially with polymerization time and might be used to characterize the polymerization extent in situ.
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- Dahlberg, Maria, et al.
(författare)
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A new chemiluminescence paradox: selective inhibition of isoluminol-amplified activity in phagocytes by peptides from annexin AI.
- 2008
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Ingår i: Luminescence : the journal of biological and chemical luminescence. - : Wiley. - 1522-7243. ; 23:3, s. 139-43
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Tidskriftsartikel (refereegranskat)abstract
- Chemiluminescence systems enhanced by either isoluminol or luminol in combination with a peroxidase are sensitive methods for the detection of reactive oxygen species (ROS) generated by phagocyte NADPH oxidase. The two amplifying substrates are structurally very similar, differing only in the position of the amino group in the aromatic ring of the molecules. This difference renders isoluminol a less lipophilic molecule that is less permeable to biological membranes. The use of isoluminol is consequently restricted to studies dealing with the secretion of oxygen metabolites. In this study we show that synthetic peptides derived from the N-terminal domain of the calcium-regulated protein annexin AI interfere with the detection of radicals in an isoluminol-amplified, but not in a luminol-amplified, system. The annexin AI-derived peptides reduce the light output with isoluminol excited by superoxide and horseradish peroxidase (HRP) in formyl-methionyl-leucyl-phenylalanine- and phorbol myristate acetate-stimulated cells, as well as by hydrogen peroxide and HRP. The precise mechanism for the inhibition is not known. The results presented strongly suggest that a reduced cellular response detected with isoluminol-amplified chemiluminescence should be confirmed with an alternative technique to determine release of superoxide anions and hydrogen peroxide.
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- Huttunen, Roope J., et al.
(författare)
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Quantitative detection of cell surface protein expression by time-resolved fluorimetry
- 2007
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Ingår i: Luminescence. - : Wiley. - 1522-7243 .- 1522-7235. ; 22:3, s. 163-170
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Tidskriftsartikel (refereegranskat)abstract
- A method is introduced for quantitative detection of cell surface protein expression. The method is based on immunocytochemistry, the use of long decay time europium(III) chelate and platinum(II) porphyrin labels, and detection of photoluminescence emission from adhered cells by time-resolved fluorimetry. After immunocytochemistry, the assay wells are evaporated to dryness and measured in the dry state. This protocol allows repeated and postponed analysis and microscopy imaging. In order to investigate the performance of the method, we chose expression of intercellular adhesion molecule-1 (ICAM-1) of endothelial cell line EAhy926 as a research target. The expression of ICAM-1 on the cells was enhanced by introduction of a cytokine, tumour necrosis factor-alpha (TNF alpha). The method gave signal: background ratios (S:B) of 20 and 9 for europium and platinum labels, respectively, whereas prompt fluorescent FITC label gave a S:13 of 3. Screening window coefficients (=Z'-factor) were >0.5 for all the three labels, thus indicating a score for an excellent screening assay. In conclusion, the method appears to be an appropriate choice for protein expression analysis, both in high-throughput screening applications, and for detailed sample investigation by fluorescent microscopy imaging. Copyright (C) 2007 John Wiley & Sons, Ltd.
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- Nurmi, Jussi, et al.
(författare)
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Time-resolved fluorometry in end-point and real-time PCR quantification of nucleic acids
- 2000
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Ingår i: Luminescence. - 1522-7235. ; 15:6, s. 381-388
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Tidskriftsartikel (refereegranskat)abstract
- Two time-resolved fluorescence-based methods for nucleic acid quantification are described and their results are compared. Both methods use an exogenous internal standard to eliminate errors arising from different steps of the assay. The first method is a competitive end-point assay, where the standard competes for the same primers with the actual target sequence, prostate-specific antigen (PSA) cDNA. The standard and target are quantified in a dual-label plate hybridization with lanthanide-labelled probes after a fixed number of PCR cycles. The second method is based on real-time monitoring of PCR and on the use of a novel homogeneous signal generation principle that relies on the use of a 5′→3′ exonucleolytic DNA polymerase and a probe labelled with an environment sensitive, stable and fluorescent lanthanide chelate. In this assay, a non-competitive, exogenous internal standard is used. Both assays have a wide linear range (50-5 × 106 and 10-5 × 107 input PSA cDNA molecules for the end-point and real-time assays, respectively) and there is a strong correlation between the results obtained with the two assays (r = 1.0). Being somewhat faster to perform, the real-time format is better suited for assays that require high throughput.
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| 15. |
- Väisänen, Ville, et al.
(författare)
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Time-resolved fluorescence imaging for quantitative histochemistry using lanthanide chelates in nanoparticles and conjugated to monoclonal antibodies
- 2000
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Ingår i: Luminescence. - 1522-7235. ; 15:6, s. 389-397
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Tidskriftsartikel (refereegranskat)abstract
- Tissue and cell examinations have a potential to produce extremely valuable information about antigen quantities in samples. Using currently available methods, a truly quantitative analysis is nearly impossible. We have previously shown that immunohistochemical (IHC) detection of prostate-specific antigen and human glandular kallikrein from prostatic tissue, together with time-resolved fluorescence imaging (TRFI), is a suitable method for obtaining quantitative data from biological samples and that the signal response is linear. In this paper we show that Eu-chelate containing particles in the nanometer range are suitable labels for quantitative IHC. Even single nanoparticle molecules can be detected by TRFI and the signals measured can be readily quantitated. The signal intensity correlates very well with the amount of bound label, and the use of nanoparticles could markedly improve the sensitivity of quantitative IHC methods. TRFI provides a powerful tool for providing quantitative data about antigens or transcripts in tissue sections or cultured cells. It is also of major importance in standardization and optimization of protocols for fixation and tissue preparation, including antigen retrieval methods.
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| 16. |
- Fäldt, Jenny, 1971, et al.
(författare)
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The phagocyte chemiluminescence paradox: luminol can act as an inhibitor of neutrophil NADPH-oxidase activity.
- 1999
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Ingår i: Luminescence : the journal of biological and chemical luminescence. - 1522-7235. ; 14:3, s. 153-60
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Tidskriftsartikel (refereegranskat)abstract
- The chemiluminescence system amplified by luminol or isoluminol is a sensitive and widely used method for determination of respiratory burst products generated by the NADPH-oxidase in phagocytes. The present study shows that luminol, but not isoluminol, can inhibit the release of oxygen metabolites generated by human neutrophil NADPH-oxidase. The difference in structure between luminol and isoluminol (rendering luminol more lipophilic than isoluminol, and thereby membrane-permeable), is suggested to determine indirectly whether or not the molecule is inhibitory. Luminol was shown to have an increased inhibitory effect after preincubation of neutrophils on a surface of aggregated IgG, suggesting that the cells can be transferred from a 'luminol-insensitive' to a 'luminol-sensitive' state. Since luminol had no inhibitory effect in a cell-free NADPH-oxidase system, it is likely that it interferes with the signal transduction pathway, leading to assembly and/or activation of the oxidase. As a consequence of the present results, showing that luminol but not isoluminol can inhibit NADPH-oxidase activity, we suggest that isoluminol is used in future studies of superoxide anion release from phagocytes.
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| 17. |
- Delroisse, J, et al.
(författare)
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Opsin-based extraocular photoreception in a luminous brittle star
- 2014
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Ingår i: Luminescence. Special Issue: Abstracts of the 18th International Symposium on Bioluminescence and Chemiluminescence, June 23–28 2014, Uppsala, Sweden (ISBC 2014). - 1522-7235. ; 29:S1, s. 65-66
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Konferensbidrag (övrigt vetenskapligt/konstnärligt)
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