Direct surface plasmon resonance immunosensor for in situ detection of benzoylecgonine, the major cocaine metabolite

Muñoz, Eva; Lorenzo-Abalde, Silvia; González-Fernández, África; Quintela, Óscar; López-Rivadulla, Manuel; Riguera, Ricardo

doi:10.1016/j.bios.2011.04.056

Cited by 31 publications

(16 citation statements)

References 43 publications

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“…The scFv-8C10 presented strong affinity to the immobilized antigen, with approximately K D of 1.8 × 10 −10 M. The fast association rate and slow dissociation rate implied strong binding between the anti-triazophos scFv-8C10 antibody and the coating antigen. However, the sensorgram for mAb showed that the whole antibody came off the immobilized surface extremely slowly and, thus, it was difficult for surface regeneration, which was also observed in a previous report [ 41 ]. In comparison to the scFv, this reaction profile of mAb could be due to the formation of stronger bivalent complexes between the mAb and the polyvalent hapten-OVA coated on the chip surface.…”

Section: Discussionsupporting

confidence: 58%

Expression and Functional Properties of an Anti-Triazophos High-Affinity Single-Chain Variable Fragment Antibody with Specific Lambda Light Chain

Liang

Xiang

et al. 2016

IJMS

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Triazophos is a widely used organophosphorous insecticide that has potentially adverse effects to organisms. In the present study, a high-affinity single-chain variable fragment (scFv) antibody with specific lambda light chain was developed for residue monitoring. First, the specific variable regions were correctly amplified from a hybridoma cell line 8C10 that secreted monoclonal antibody (mAb) against triazophos. The regions were then assembled as scFv via splicing by overlap extension polymerase chain reaction. Subsequently, the recombinant anti-triazophos scFv-8C10 was successfully expressed in Escherichia coli strain HB2151 in soluble form, purified through immobilized metal ion affinity chromatography, and verified via Western blot and peptide mass fingerprinting analyses. Afterward, an indirect competitive enzyme-linked immunosorbent assay was established based on the purified anti-triazophos scFv-8C10 antibody. The assay exhibited properties similar to those based on the parent mAb, with a high sensitivity (IC50 of 1.73 ng/mL) to triazophos and no cross reaction for other organophosphorus pesticides; it was reliable in detecting triazophos residues in spiked water samples. Moreover, kinetic measurement using a surface plasmon resonance biosensor indicated that the purified scFv-8C10 antibody had a high affinity of 1.8 × 10−10 M and exhibited good binding stability. Results indicated that the recombinant high-affinity scFv-8C10 antibody was an effective detection material that would be promising for monitoring triazophos residues in environment samples.

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Section: Discussionsupporting

confidence: 58%

Expression and Functional Properties of an Anti-Triazophos High-Affinity Single-Chain Variable Fragment Antibody with Specific Lambda Light Chain

Liang

Xiang

et al. 2016

IJMS

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“…Covalent immobilization strategy was selected as reference of a standard and commonly used procedure that generally leads to randomly oriented layer of antibodies. It inherently generates high biosurface stability and allows the control of the packing density although it usually requires high concentration of antibody, between 0.1 and 1 mg/mL [21,22]. Protein G (or Protein A)-mediated immobilization strategy has been extensively used in the biosensing field [23,24].…”

Section: Discussionmentioning

confidence: 99%

Direct Detection of Protein Biomarkers in Human Fluids Using Site-Specific Antibody Immobilization Strategies

Soler

Estévez

Álvarez

et al. 2014

Sensors

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Design of an optimal surface biofunctionalization still remains an important challenge for the application of biosensors in clinical practice and therapeutic follow-up. Optical biosensors offer real-time monitoring and highly sensitive label-free analysis, along with great potential to be transferred to portable devices. When applied in direct immunoassays, their analytical features depend strongly on the antibody immobilization strategy. A strategy for correct immobilization of antibodies based on the use of ProLinker™ has been evaluated and optimized in terms of sensitivity, selectivity, stability and reproducibility. Special effort has been focused on avoiding antibody manipulation, preventing nonspecific adsorption and obtaining a robust biosurface with regeneration capabilities. ProLinker™-based approach has demonstrated to fulfill those crucial requirements and, in combination with PEG-derivative compounds, has shown encouraging results for direct detection in biological fluids, such as pure urine or diluted serum. Furthermore, we have implemented the ProLinker™ strategy to a novel nanoplasmonic-based biosensor resulting in promising advantages for its application in clinical and biomedical diagnosis.

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“…Because of the above reasons, the most‐used ligands for COC detection are antibodies. Conventional ELISA (enzyme‐linked immunosorbent analysis) 5–7, 23 and immunosensors 24–28 both have high sensitivity (smaller LODs are around 0.1–10 ng mL −1 COC) and specifity due to antibodies. Most of these ELISAs, which are used for testings of drug of abuse, only detect positive BZE concentrations with a typical cutoff value of 300 ng mL −1 , instead of free unhydrolyzed COC.…”

Section: Introductionmentioning

confidence: 99%

“…Usually, complete extraction or cleanup of the biological samples is necessary in these ELISA methods due to the complex biological matrices interfering with antibodies. Immunosensors for COC and their metabolites include surface‐plasmon resonance 24, piezoelectric 25, surface acoustic wave 26, fluorescence fiber‐optic 27, or porous silicon photonic 28, with lower LODs to values of 4 ng mL −1 , but hardly electrochemical immunosensors despite the inherent sensitivity in measuring currents.…”

Section: Introductionmentioning

confidence: 99%

A Multi‐electrochemical Competitive Immunosensor for Sensitive Cocaine Determination in Biological Samples

Vidal

Bertolín

Bonel³

et al. 2015

Electroanalysis

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[a] 1Introduction Cocaine (COC) is ap owerfully addictive alkaloidp resent on the leaves of the coca plant (erythroxylum coca, erythroxylumn ovogranatense)a nd is one of the most worldwide used illicit drugs [1].I ti sastrong centraln ervous system stimulantt hat increases the levels of the neurotransmitter dopamine by blocking reuptakes at nerve synapses,t hereby prolonging its action.Following administration, COC is rapidly metabolized by enzymatic hydrolysis into two majorm etabolites benzoylecgonine( BZE) and ecgonine methyl-ester (EME) [2].C OC concentrations in plasma depend from the routes of intake, and maximum values vary in ar ange 50-2000 ng mL À1 ,i natime of about 30-120 min. or greater depending on the route [3].N evertheless,m etabolites may be identified up to 144 h. Metabolite:cocaine ratios increase after COC administration, and might prove useful for interpreting times of last use in pharmacological and pharmacokineticss tudies [4].C OC is eliminated unchanged in the urine in 1-9 %p ercentage,w ith ah igher proportion in acid urine [2,3].T he duration of unmetabolized cocaine in urine can be up to 6-8 hours,so that the pharmacologicald etection of COC also reports av ery recent COC use [3].T he initial screening cutoff levels of COC in serum and urine are usually considered 300 ng mL À1 for cocaine and its metabolites [2,3]. Fort he samei ndividual, COC concentration in saliva is approximately three-fold of that foundi np lasma and over five-fold in urine.I na ddition, COC elimination halflife is lower in saliva than in urine or blood, which makes oral fluid analysis suitable for determination of very recent use providing unequivocal screen results within minutes [4].T he cutoff level on saliva drug tests for cocaine is normally 20 ng mLTheb iological detection of drugu se involvesascreening test, which if positive,i sf ollowed by ac onfirmatory test. Screeninga nalysis of COC in av ariety of samples (e.g. urine,b lood,p lasma, hair, saliva, sweat, meconium, cocaine powder or waste water) use commonly enzyme immunoassays (EIA), e.g. ELISA (enzyme-linked immunosorbent analysis) [5][6][7],w hile liquid chromatography-(tandem) masss pectrometry( LC-MS/MS) and gas chromatography-mass spectrometry (GC-MS) are the usual methods of choice for confirmation of presumptive positive results from the initial screeninga nalysis [8].T ypically,C OC is extracted in an organic solventf or chromatographic analysis,f ollowed by af orm of sample clean-up (e.g. centrifugation or filtration), preconcentration, and purification of the extractedt arget due to matrix effects [8].G C-MS also require complicated chemical derivatization procedures of the COC molecules [5,8].S uch timeexpensives amplep retreatment procedures can be avoided with immunochemical analysis simply by diluting the biological sample in an aqueous buffer beforethe analysis [5].T he reported limits of detection (LODs) of the GC-MS and LC-MS/MS methods are in the range1 .2 to Abstract:Am ulti-electrochemical competitive immunosensor for the rapi...

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Direct surface plasmon resonance immunosensor for in situ detection of benzoylecgonine, the major cocaine metabolite

Cited by 31 publications

References 43 publications

Expression and Functional Properties of an Anti-Triazophos High-Affinity Single-Chain Variable Fragment Antibody with Specific Lambda Light Chain

Expression and Functional Properties of an Anti-Triazophos High-Affinity Single-Chain Variable Fragment Antibody with Specific Lambda Light Chain

Direct Detection of Protein Biomarkers in Human Fluids Using Site-Specific Antibody Immobilization Strategies

A Multi‐electrochemical Competitive Immunosensor for Sensitive Cocaine Determination in Biological Samples

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