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Piezoelectric immunosensors for urine specimens of Chlamydia trachomatis employing quartz crystal microbalance microgravimetric analyses
ABST~CTS
Pharmacological evidence suggests that the neurotransmitter released by vertebrate photoreceptors is L-glutamate. To confirm and study this release, we developed a, glutamate-sensitive microelectrode (10-30 ~tm tip diameter) consisting of a glass-encased Pt-wire whose exposed extremity was coated with a polypyrrole layer containing glutamate oxidase. Glutamate was detected amperometrically as hydrogen peroxide produced from oxidized L-glutamate (Pt-wire polarized at 500 mV against Ag/AgC1). Exposure to 100 jaM L- glutamate caused an increase in microelectrode current of about 50 pA; glutamate concentration changes down to about 1 ~tM were detectable. Electrode sensitivity was 100-fold greater for L- glutamate than for other major amino-acids. However, oxidizable compounds (eg catecholamines) crossing the polypyrrole layer were also detected. We used this microelectrode to measure activitydependent glutamate release from single presynaptic terminals of solitary photoreceptors from salamander retina; various strategies were followed, but we failed to detect any release of glutamate. In contrast, on a slice of insect retina where non-vesicular release of glutamate occurs, we measured a glutamate concentration of about 10 ~M close to the surface of the retina. Possible reasons for our failure to detect glutamate released from vertebrate photoreceptors are discussed.
Biosensors & Bioelectronics Vol. 13 No. i (1998)
not in the PNA. Sputter-initiated resonance ionization microprobe analysis was used to detect the presence of enriched tin isotope-labelled DNA and of phosphorus in natural DNA as a means to identify the presence of DNA after hybridization to oligodeoxynucleotides (ODNs) or PNAs, respectively, immobilized on a biosensor chip. The data clearly demonstrate that excellent discrimination between complementary and noncomplementary sequences can be obtained during hybridization of DNA to either ODNs or PNAs. The capability to detect different enriched stable isotope-labelled DNAs simultaneously allows high degrees of multiplexing which may be very advantageous for hybridization kinetics studies in complex systems, as well as significantly increasing the speed of analysis. Alternatively, by using natural DNA with PNA biosensor chips, discrimination for single-point mutation could be increased because of improved hybridization kinetics and direct analysis of genomic DNA may become possible without amplification, Both methods have the potential to provide a rapid method for DNA/RNA sequencing, diagnostics, and mapping. Piezoelectric immunosensors for urine specimens of Chlamydia trachomatis employing quartz crystal microbalance microgravimetric analyses BenDov, I; Willner, I & Zisman, E
Analysis of biosensor chips for identification of nucleic acids
ANALYTICAL CHEMISTRY (1997) 69(17): 3506-3512
Arlinghaus, HF; Kwoka, MN & Jacobson, KB
The assembly of a biosensor for Chlamydia trachomatis based on the microgravimetric quartz crystal microbalance (QCM) analysis of the bacteria association to an antibody-functionalized electrode is described. The sensing interfaces consist of a primary cystamine monolayer assembled onto Au electrodes associated with the quartz crystal. The monolayer is further modified with sulfosuccinylimidyl 4-(p-maleimidophenyl) butyrate (sulfo-SMPB) and the goat IgG-anti-mouse IgG Fc-specific Ab or the fragmented F(ab')(2) anti-mouse IgG Ab that act
ANALYTICAL CHEMISTRY (1997) 69(18): 3747-3753 Two novel DNA-sequencing methods are described that use DNA hybridization biosensor chips. These two techniques involve either labelling the free nucleic acid .with enriched stable isotopes or hybridizing DNA without labels to immobilized peptide nucleic acid (PNA) and detecting the phosphorus present in the DNA but
iv
Biosensors & Bioelectronics VoL 13 No. 1 (1998)
as sublayers for the association of the sensor-active anti-C, trachomatis LPS-Ab. Bacteria in the concentration range from 260 ng/mL to 7.8 lag/mL are sensed by the functionalized crystals. The association of C. trachomatis to the sensing interface can be confirmed and amplified via interaction of the crystal with various anti-C, trachomatis antibodies, Urine-pretreated functionalized quartz crystals are applied in the analysis of C. trachomatis in urine samples. The sensitivity limits of the electrodes for sensing the bacteria in urine samples corresponds to similar to 260 ng.mL ~. The functionalized crystals assembled via association of anti-C, trachomatis LPS-Ab to the fragmented F(ab')(2) anti-mouse IgG Ab reveal long-term stability upon storage at 4 degrees C.
Real-time monitoring of peptide-surface and peptide-antibody interaction by means of reflectometry and surface plasmon resonance Loomans, EEMG; Beumer, TAM; Damen, KCS; Bakker, MA & Schielen, WJG JOURNAL OF COLLOID AND INTERFACE SCIENCE (1997) 192(1): 238-249 The performance of immunodiagnostic assays such as ELISA is governed by many different factors. Reflectometry was used to monitor peptide adsorption and the resulting antibody binding activity on a polystyrene surface, Surface plasmon resonance was used to analyze affinity and kinetic parameters of the (immobilized) peptide-antibody interaction. We demonstrate the capability of both the reflectometer and the BIAcore instrument to determine these immunoassay factors independently. When peptidic antigens other than the parent protein antigen were applied, reduced antibody binding activity (10 times lower) and faster dissociation (100 times faster') rather than poor adsorption proved to be the critical factors determining immune reactivity. When the peptides were modified chemically or when their molecular size
0956-5663/98/$19.00©1998 Elsevier Science S.A.
ABSTRACTS
was increased, antibody binding activity as well as affinity could be improved or even restored.
Fiber optic genosensor for specific determination of femtomolar DNA oligomers Kleinjung, F; Bier, FF; Warsinke, A & Scheller, FW ANALYTICA CHIMICA ACTA (1997) 350(12): 51-58 The binding of DNA oligonucleotides to immobilized DNA-targets using a fiber optic fluorescence sensor is demonstrated. 13mer oligonucleotides were attached to the core of a multimode fiber. The complementary sequence was detected by the use of a fluorescent double strand specific DNA ligand (YOYO and PicoGreen). The evanescent field was employed to distinguish between bound and not bound species. The template DNA-oligomer was immobilized either by direct coupling to the activated sensor surface or using the avidin-biotin bridge. Single base mismatches in the target sequence were detected; and a detection limit of the sensor of 30fM (3.2 amol) was found for the matching target.
v
Pharmacological evidence suggests that the neurotransmitter released by vertebrate photoreceptors is L-glutamate. To confirm and study this release, we developed a, glutamate-sensitive microelectrode (10-30 ~tm tip diameter) consisting of a glass-encased Pt-wire whose exposed extremity was coated with a polypyrrole layer containing glutamate oxidase. Glutamate was detected amperometrically as hydrogen peroxide produced from oxidized L-glutamate (Pt-wire polarized at 500 mV against Ag/AgC1). Exposure to 100 jaM L- glutamate caused an increase in microelectrode current of about 50 pA; glutamate concentration changes down to about 1 ~tM were detectable. Electrode sensitivity was 100-fold greater for L- glutamate than for other major amino-acids. However, oxidizable compounds (eg catecholamines) crossing the polypyrrole layer were also detected. We used this microelectrode to measure activitydependent glutamate release from single presynaptic terminals of solitary photoreceptors from salamander retina; various strategies were followed, but we failed to detect any release of glutamate. In contrast, on a slice of insect retina where non-vesicular release of glutamate occurs, we measured a glutamate concentration of about 10 ~M close to the surface of the retina. Possible reasons for our failure to detect glutamate released from vertebrate photoreceptors are discussed.
Biosensors & Bioelectronics Vol. 13 No. i (1998)
not in the PNA. Sputter-initiated resonance ionization microprobe analysis was used to detect the presence of enriched tin isotope-labelled DNA and of phosphorus in natural DNA as a means to identify the presence of DNA after hybridization to oligodeoxynucleotides (ODNs) or PNAs, respectively, immobilized on a biosensor chip. The data clearly demonstrate that excellent discrimination between complementary and noncomplementary sequences can be obtained during hybridization of DNA to either ODNs or PNAs. The capability to detect different enriched stable isotope-labelled DNAs simultaneously allows high degrees of multiplexing which may be very advantageous for hybridization kinetics studies in complex systems, as well as significantly increasing the speed of analysis. Alternatively, by using natural DNA with PNA biosensor chips, discrimination for single-point mutation could be increased because of improved hybridization kinetics and direct analysis of genomic DNA may become possible without amplification, Both methods have the potential to provide a rapid method for DNA/RNA sequencing, diagnostics, and mapping. Piezoelectric immunosensors for urine specimens of Chlamydia trachomatis employing quartz crystal microbalance microgravimetric analyses BenDov, I; Willner, I & Zisman, E
Analysis of biosensor chips for identification of nucleic acids
ANALYTICAL CHEMISTRY (1997) 69(17): 3506-3512
Arlinghaus, HF; Kwoka, MN & Jacobson, KB
The assembly of a biosensor for Chlamydia trachomatis based on the microgravimetric quartz crystal microbalance (QCM) analysis of the bacteria association to an antibody-functionalized electrode is described. The sensing interfaces consist of a primary cystamine monolayer assembled onto Au electrodes associated with the quartz crystal. The monolayer is further modified with sulfosuccinylimidyl 4-(p-maleimidophenyl) butyrate (sulfo-SMPB) and the goat IgG-anti-mouse IgG Fc-specific Ab or the fragmented F(ab')(2) anti-mouse IgG Ab that act
ANALYTICAL CHEMISTRY (1997) 69(18): 3747-3753 Two novel DNA-sequencing methods are described that use DNA hybridization biosensor chips. These two techniques involve either labelling the free nucleic acid .with enriched stable isotopes or hybridizing DNA without labels to immobilized peptide nucleic acid (PNA) and detecting the phosphorus present in the DNA but
iv
Biosensors & Bioelectronics VoL 13 No. 1 (1998)
as sublayers for the association of the sensor-active anti-C, trachomatis LPS-Ab. Bacteria in the concentration range from 260 ng/mL to 7.8 lag/mL are sensed by the functionalized crystals. The association of C. trachomatis to the sensing interface can be confirmed and amplified via interaction of the crystal with various anti-C, trachomatis antibodies, Urine-pretreated functionalized quartz crystals are applied in the analysis of C. trachomatis in urine samples. The sensitivity limits of the electrodes for sensing the bacteria in urine samples corresponds to similar to 260 ng.mL ~. The functionalized crystals assembled via association of anti-C, trachomatis LPS-Ab to the fragmented F(ab')(2) anti-mouse IgG Ab reveal long-term stability upon storage at 4 degrees C.
Real-time monitoring of peptide-surface and peptide-antibody interaction by means of reflectometry and surface plasmon resonance Loomans, EEMG; Beumer, TAM; Damen, KCS; Bakker, MA & Schielen, WJG JOURNAL OF COLLOID AND INTERFACE SCIENCE (1997) 192(1): 238-249 The performance of immunodiagnostic assays such as ELISA is governed by many different factors. Reflectometry was used to monitor peptide adsorption and the resulting antibody binding activity on a polystyrene surface, Surface plasmon resonance was used to analyze affinity and kinetic parameters of the (immobilized) peptide-antibody interaction. We demonstrate the capability of both the reflectometer and the BIAcore instrument to determine these immunoassay factors independently. When peptidic antigens other than the parent protein antigen were applied, reduced antibody binding activity (10 times lower) and faster dissociation (100 times faster') rather than poor adsorption proved to be the critical factors determining immune reactivity. When the peptides were modified chemically or when their molecular size
0956-5663/98/$19.00©1998 Elsevier Science S.A.
ABSTRACTS
was increased, antibody binding activity as well as affinity could be improved or even restored.
Fiber optic genosensor for specific determination of femtomolar DNA oligomers Kleinjung, F; Bier, FF; Warsinke, A & Scheller, FW ANALYTICA CHIMICA ACTA (1997) 350(12): 51-58 The binding of DNA oligonucleotides to immobilized DNA-targets using a fiber optic fluorescence sensor is demonstrated. 13mer oligonucleotides were attached to the core of a multimode fiber. The complementary sequence was detected by the use of a fluorescent double strand specific DNA ligand (YOYO and PicoGreen). The evanescent field was employed to distinguish between bound and not bound species. The template DNA-oligomer was immobilized either by direct coupling to the activated sensor surface or using the avidin-biotin bridge. Single base mismatches in the target sequence were detected; and a detection limit of the sensor of 30fM (3.2 amol) was found for the matching target.
v