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Biomedical subjects

E Livaniou

Publications and source records attributed to E Livaniou.

27 records · Page 2Linked to original sources
PubMed1992

A thymosin beta 4 ELISA using an antibody against the N terminal fragment thymosin beta 4 [1-14].

A thymosin beta 4 ELISA was developed in which thymosin beta 4, absorbed on microwells, competed with thymosin beta 4 in solution for the binding sites of an anti-thymosin beta 4 antibody. The antibody molecules finally immobilized on the microwells were detected using a goat anti-rabbit immunoglobulin/horseradish peroxidase conjugate in combination with the substrate 2,2'-azino-bis-(3-ethylbenzthiazoline-6-sulfonic acid) diammonium salt, and measuring the relevant optical density values. Anti-thymosin beta 4 antibodies were raised in rabbits against intact thymosin beta 4 as well as against selected fragments of the peptide, i.e., the N terminal fragments thymosin beta 4[1-14] and thymosin beta 4[1-11]. The antibody against thymosin beta 4[1-14] was used in the thymosin beta 4 ELISA, because it showed minimal cross-reactivity (0.1%) with the highly homologous peptide thymosin beta 9 as well as exhibiting the highest titre. The ELISA procedure developed, apart from showing a minimal cross-reaction with thymosin beta 9, was fast, easy to perform and exhibited good assay characteristics.

Animals↗
PubMed1992

Preparation of the N-(4'-hydroxy-[3'-125I]iodophenethyl)-6-(4-O-diethylstilbestryl)hexanam ide for diethylstilbestrol radioimmunoassays.

The synthesis of a radioiodinated diethylstilbestrol (DES) derivative is described. This derivative was prepared by coupling the previously synthesized active ester of 6-(4-O-diethylstilbestryl)hexanoic acid with mono-[125I]iodotyramine in dry tetrahydrofuran (20 to 22 C, 16 hours). The mono-[125I]iodotyramine was prepared using a chloramine-T method and purified by paper electrophoresis. The final product, N-(4'-hydroxy-[3'-125I]iodophenethyl)-6-(4-O- diethylstilbestryl)hexanamide, was separated by thin-layer chromatography (cyclohexane/ethanol/NH4OH 2.5 N/acetone; 40:50:5:20, v/v/v/v); it was stable for 2 months in ethanol at 4 C and had a specific activity higher than 540 Ci/mmol. The [125I]DES amide synthesized was found to retain the immunoreactivity of DES, since it competed with [3H]DES or DES in an in vitro radioimmunoassay system for the binding sites of a rabbit anti-DES antibody; thus, it seems to be capable of replacing the tritiated tracer used so far in DES radioimmunoassays.

Binding, Competitive↗
PubMed1991

Biotinidase radioassay using an 125I-biotin derivative, avidin, and polyethylene glycol reagents.

A radioassay for determining biotinidase activity in human serum was developed, using N-[beta-(4-OH-3-125I-phenyl)ethyl]-biotinamide in combination with biocytin as the substrate, avidin as a binding protein, and polyethylene glycol as a separation reagent. The gamma-emitting 125I-biotinamide (= tracer) was synthesized by coupling (pH 8.5, 20-22 degrees C, 90 min) N-hydroxysuccinimidobiotin to 125I-tyramine. Using polyethylene glycol as a separation reagent, it was possible to eliminate several problems that were encountered when other separation reagents were used. Biotinidase activity was evaluated following the cleavage of the 125I-biotinamide and expressed in fmol of tracer cleaved.min-1.ml-1 in the presence of 9 nmol of biocytin. Under the conditions used, the time response of the assay was linear up to 3 h. The method is simple to perform, more sensitive than the previously described methods, and reproducible (intra- and interassay CVs of 4.9 and 10.2%, respectively) and allows the simultaneous handling of more than 100 samples in less than 3 h.

Amidohydrolases↗
PubMed1991

Isolation of mono- and di-iodine 125 tyramines for conjugation labelling.

Here we describe a method for isolating mono- and di-iodine 125 tyramines from a crude tyramine radio-iodination reaction mixture. Tyramine was radio-iodinated according to a modified chloramine-T method. Mono- and diforms were separated from each other, as well as from free 125I and unreacted tyramine, by means of paper electrophoresis or thin-layer chromatography. Both 125I-tyramine derivatives finally obtained were pure and had high specific activities, which averaged 2500 Ci/mmol for mono-125I-tyramine and 5000 Ci/mmol for di-125I-tyramine. Due to their characteristics, these derivatives can be used for labelling various substances indirectly and preparing in vivo and in vitro tracers of high purity and specific activity, when conjugation labelling is required.

Iodine Radioisotopes↗
PubMed1991

Plasma biotin levels in neonates.

Plasma biotin levels were measured by a new, very sensitive radioligand assay, in 47 normal full-term infants, receiving breast milk and/or different formulas, during the first 3 weeks of life. Exclusively breast fed infants and infants fed with a formula containing 11 micrograms/l biotin had plasma biotin levels of 538 +/- 78 and 502 +/- 117 ng/l, respectively, during the 1st week of life, and 321 +/- 48 and 503 +/- 153 ng/l, respectively, during the 3rd week of life. Infants fed with a formula very high in biotin (300 micrograms/l) had extremely high plasma biotin levels during the 1st week of life (13,741 +/- 659 ng/l) while during the 2nd week of life they showed a tendency towards decreasing plasma biotin levels. These changes of plasma biotin levels during the first weeks of life indicate that the in vivo kinetics of biotin is changing during this period.

Aging↗
PubMed1987

Biotin radioligand assay with an 125I-labeled biotin derivative, avidin, and avidin double-antibody reagents.

We describe a new radioligand assay for determining biotin in biological fluids by using a mixture of N-[beta-(4-OH-3-125I-phenyl)ethyl]- and N-[beta-(4-OH-3,5-di-125I-phenyl)ethyl]biotinamides as radiotracer, avidin as a binding protein, and an avidin double-antibody as a separation reagent. The radiotracer is synthesized by coupling (at pH 8.5, 20-22 degrees C, 90 min) N-hydroxysuccinimidobiotin to radioiodinated tyramine. The assay curve is linear and the assay itself is sensitive (less than 10 ng/L), reproducible (intra- and interassay CVs 4.1% and 7.0%, respectively), and allows the simultaneous handling of more than 100 samples in less than 4 h. Serum samples from apparently normal subjects contained 100-840 ng of biotin per liter (mean 340 ng/L). Pregnant women had low concentrations of biotin (100-300 ng/L) in their serum. Patients undergoing chronic hemodialysis treatment showed high concentrations (0.5-3.0 micrograms/L), which may be ascribable to the inability of avidin, which was used as the assay binding protein, to distinguish biotin from biotinyl derivatives with an intact ureido ring.

Antibodies↗
PubMed1987

Radioiodinated biotin derivatives for in vitro radioassays.

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 degrees 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-[beta-(4-OH-3-125I-phenyl)ethyl] and the N-[beta-(4-OH-3,5-di 125I-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 complete 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 high-greater than 350 Ci/mmol-and they were stable for 2 mo at 4 degrees C.

Avidin↗