Supp. Information JMC 2013 (original) (raw)

Grubb et al. Clin Chem Lab Med 2010

The IFCC Working Group for the Standardisation of Cystatin C (WG-SCC), in collaboration with the Institute for Reference Materials and Measurements (IRMM), announces the availability of the new certified reference material ERM-DA471/IFCC. The material was characterised using a pure protein primary reference preparation (PRP) as calibrant. The PRP was prepared from recombinant cystatin C, and its concentration measured using dry mass determination. The characterisation of ERM-DA471/IFCC was performed by particle enhanced immuno-nephelometry, particle enhanced immuno-turbidimetry, and enzyme amplified single radial immuno-diffusion. The certified cystatin C mass concentration in ERM-DA471/IFCC, if reconstituted according to the specified procedure, is 5.48 mg/L, the expanded uncertainty (ks2) being 0.15 mg/L. Clin Chem Lab Med 2010;48:1619-21.

Development of a versatile biotinylated material based on SU-8

Journal of Materials Chemistry B, 2013

The negative epoxy-based SU-8 photoresist has a wide variety of applications within the semiconductor industry, photonics and lab-on-a-chip devices, and it is emerging as an alternative to silicon-based devices for sensing purposes. In the present work, biotinylation of the SU-8 polymer surface promoted by light is reported. As a result, a novel, effective, and low-cost material, focusing on the immobilization of bioreceptors and consequent biosensing, is developed. This material allows the spatial discrimination depending on the irradiation of desired areas. The most salient feature is that the photobiotin may be directly incorporated into the SU-8 curing process, consequently reducing time and cost. The potential use of this substrate is demonstrated by the immunoanalytical detection of the synthetic steroid gestrinone, showing excellent performances. Moreover, the naked eye biodetection due to the transparent SU-8 substrate, and simple instrumental quantification are additional advantages.

Chapter 8. Labelling of biotin with 188Re

2015

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A spectrophotometric assay for nanogram quantities of biotin and avidin

Journal of Biochemical and Biophysical Methods, 1986

Parameters and conditions of an enzyme based assay for biotin and avidin are presented. Biotinylated glucose-6-phosphate dehydrogenase when complexed with avidin becomes inactivated. Thus it was possible to construct a competitive assay system for biotin. The assay is sensitive between 100-500 ng/ml and could detect as little as 10 ng in 0.1 ml with a between run error of 2.4%. It requires a 60 rain incubation at 21°C and 5 rain to assay. The avidin assay, based on the degree of inactivation of biotinylated-glucose-6-phosphate dehydrogenase in relation to the concentration of avidin, could detect as little as 0.25 ng in 0.1 ml or 2.5 ng/ml with an assay time of 10 rain with a between run error of 3.9%. Both assays are rapid with significant improvements over other non-isotopic methods in sensitivity and comparable to radioisotopic methods in sensitivity with the added advantage of ease of method.

Fluorometric assay for quantitation of biotin covalently attached to proteins and nucleic acids.

As a component of the (strept)avidin affinity system, biotin is often covalently linked to proteins or nucleic acids. We describe here a microplate-based high-throughput fluorometric assay for biotin linked to either proteins or nucleic acids based on fluorescence resonance energy transfer (FRET). This assay utilizes a complex of Alexa Fluoro 488 dye-labeled avidin with a quencher dye, 2-(4'-hydroxyazobenzene) benzoic acid (HABA), occupying the biotin binding sites of the avidin. In the absence of biotin, HABA quenches the fluorescence emission of the Alexa Fluor 488 dyes via FRET HABA is displaced when biotin binds to the Alexa Fluor 488 dye-labeled avidin, resulting in decreased FRET efficiency. This mechanism results in an increase in fluorescence intensity directly related to the amount of biotin present in the sample. The assay is able to detect as little as 4 pmol biotin in a 0.1 mL volume within 15 min of adding sample to the reagent, with a Z-factor > 0.9.

68 Ga-Labeling of Biotin Analogues and their Characterization

Bioconjugate Chemistry, 2009

Biotin-and 68 Ga-based tracers have been suggested as tools that could be used to monitor the survival of avidincoated islets of Langerhans isolated from pancreas and used in transplantation, i.e., to liver. Three biotin analogues with various alkyl and poly(ethylene glycol) (PEG) chains coupled to DOTA were synthesized and labeled with 68 Ga. The 68 Ga labeling was studied at room temperature as well as elevated temperature using either conventional or microwave heating. Radioactivity incorporation reached 95% within 5 and 2 min using the, respectively, conventional and microwave heating modes. The specific activity of the tracers was improved by preconcentration and purification of the generator eluate. The binding of the labeled and nonlabeled conjugates to avidin in solution was compared to the binding of native biotin. All compounds maintained good affinity for avidin, though introducing the linkers and chelator, especially the PEG-groups, somewhat decreased the binding affinity. The extent of binding of the labeled compounds to avidin was 54-91% after 5 min. Blocking experiments were performed confirming the specificity of the binding of biotin analogues to avidin. The stability of the three labeled compounds in human serum was studied. The stability of the biotin analogue 8 (65% within 30 min) and avidin-biotin complex (80% within 120 min) might be sufficient for the monitoring of the islets of Langerhans. The tracers will be evaluated in in Vitro experiments of avidin-coated islets of Langerhans and in transplantation models in ViVo.

Radioiodinated Biotin Derivatives for In Vitro Radioassays

Journal of Nuclear Medicine, 1987

The synthesis of two radioiodinated biotin derivatives with the biotin-ureido group intact is described. This synthesis was performed by coupling (pH 8.5, 20-22°C, 90 min) N-hydroxysuccinimidobiotin to tyramine, which was radioiodinated prior to this using a modified chloramine-T method. Two derivatives were produced, the N-[/3-(4-OH-3-125lphenyl)ethyl] and the N-[/i-{4-OH-3,5-di125l-phenyl)ethyl] biotin amides, depending on the amount of tyramine used in the radioiodination reaction. The final products were separated by thin layer chromatography (n-butanol: 2N NH4OH: ethanol,3:1:1, v/v/v). The radioiodinated derivatives that were synthesized or their resulting mixture were found to compete with biotin for the avidin-binding sites; thus, they were capable of being used as tracers in biotin radioassays. The specific activity of their mixture was highâ€">350 Ci/mmolâ€"and they were stable for 2 mo at 4°C.

Analytical techniques for determining biotin

… of Chromatography A, 2000

Biotin is a vitamin of the B-complex, which plays an important biochemical role in every living cell. In the recent years, the interest in this vitamin has been rekindled, mainly due to its association with serious human disorders, such as the inherited syndrome multiple carboxylase deficiency, which can be successfully treated with biotin administration. Diagnosis of biotin deficiency as well as monitoring of biotin levels in biological fluids of patients receiving biotin treatment is crucial. Equally important is the determination of biotin levels in pharmaceutical preparations as well as in food and food supplement products, which constitute the main source of biotin in humans. Several analytical methods for measuring biotin in various samples, e.g. human fluids, pharmaceutical formulations, food material etc., have been reported in the literature. In this review, the most representative of these methods are presented, and their characteristics are evaluated.

Photocleavable biotin derivatives: a versatile approach for the isolation of biomolecules

Proceedings of the National Academy of Sciences, 1995

While the strong biotin-avidin interaction has been widely used for the detection of biomolecules, its irreversibility complicates their isolation. We report the synthesis of a photocleavable biotin derivative (PCB) which eliminates many limitations of existing methods. This reagent contains a biotin moiety linked through a spacer arm to a photocleavable moiety, which reacts selectively with primary amino groups on any substrate. In experiments using [leucine]enkephalin as a model substrate, we show that PCB retains its high affinity toward avidin/streptavidin and allows rapid (<5 min) and efficient (>99%o) photorelease of the substrate in a completely unaltered form. Photocleavable biotins should be useful in numerous applications involving the isolation of proteins, nucleic acids, lipids, and cells.

Sulpho-N-hydroxysuccinimide activated long chain biotin

Journal of Immunological Methods, 1991

Biotinamidohexanoic acid N-hydroxysulphosuccinimide ester (N-hydroxysulphosuccinimide activated long chain biotin; sulpho-NHS-LC-biotin) has become an invaluable tool for the biotinylation of protein despite the absence of data concerning its stability and reaction velocity. A convenient, rapid and sensitive assay for this compound has been developed based on the sulpho-NHS-LC-biotin mediated biotinylation of bovine serum albumin following adsorption to the wells of a microtitre plate. Bound biotin was visualized by the sequential use of streptavidin and biotinylated horseradish peroxidase. This assay was used for the determination of the stability of sulpho-NHS-LC-biotin in aqueous solution of different pH values. Hydrolysis half lives were below 15 min at pH values above 8.0, but at pH values below 6.5 they exceeded 2 h. It is suggested, therefore, that biotinylations should be performed with sulpho-NHS-LC-biotin taken from a stock solution, prepared at pH values between 3.0 and 5.8. Reaction velocities with primary amino groups wc~e also investigated by means of this ELISA procedure. As expected, biotinylation proceeds faster at pH 8.0 as compared to 7.2, but the increased reaction rate does not compensate for the decreased hydrolysis half life at the higher pH value. Thus, bioli~ylation with sulpho-NHS-LC-biotin at near neutral pH values air, pears to be optimal.