A solid phase radioimmunoassay for triiodothyronine using antibody-immobilized serum albumin microspheres

A solid phase radioimmunoassay for triiodothyronine (T₃) has been developed using antibody-immobilized serum albumin microspheres. Antibody albumin microspheres were prepared using a spinning disc aerosol generator. The low density of the antibody-albumin microspheres gives greater mobility for the particles there by ensuring better kinetics to the antigen-antibody reaction. The assay has a single incubation of one hour at 37° C and the separation of the antigen-antibody complex is accomplished by centrifugation. The sensitivity of this assay is 0.3 ng/cm³ and has a range of 0.3–4 ng/cm³.     

Solid phase radioimmunoassay of triiodothyronine (T3) using antibody coupled carboxymethylcellulose powder

The paper describes a solid-phase radioimmunoassay for triiodothyronine (T₃) using antibody coupled to carboxymethylcellulose powder. The free carboxylic acid groups of cellulose are covalently coupled to the amino groups of the antibody using 1-ethyl-3-(3-dimethylamino-propyl) carbodiimide hydrochloride. Immobilized antibody thus prepared was used in a solid-phase radioimmunoassay of T₃. The assay has a sensitivity of 0.18 g/l, and a range of 0.18–4 g/l. Satisfactory correlation was obtained when this assay was compared with a T₃ assay based on dextran coated charcoal separation system (Y=0.95X+0.15 g/L; r=0.98).     

A rapid magnetizable solid phase radioimmunoassay kit for human placental lactogen

The radioimmunoessay of human placental lactogen (HPL) with separation of antibody bound and free hormone was achieved by the magnetizable solid phase coupled to antibody. The precision, accuracy, sensitivity and specificity of the method has been carefully checked in this study and the procedure of the assay was performed at room temperature. The above parameters were evaluated by recovery test (99%); within assays (4%), between assays (5%), sensitivity (0.04 g/ml) and there was no obvious cross reactivity with human growth hormone (hGH) and human chorionic gonadotropin (hCG).     

Development of solid phase radioimmunoassay system using new polymeric magnetic micro-spheres

Magnetic particles were locally prepared by co-precipitation of Fe²⁺ and Fe³⁺ in an ammonia solution. The prepared microsphere were grafted with polyacrylamide acrylic acid by using gamma irradiation polymerization in presence of MBA as a cross linker. AFP antibody was immobilized on these beads and used as a solid phase in radioimmunoassay technique. The immunoreactivity of the developed assay was found to be influenced by different factors such as solid phase volume, incubation time, incubation temperature and storage period. A comparative study was performed between the developed assay system and others two ones. The maximum binding percent attained the value of 19.5% while the sensitivity was observed to be 1.3 IU/mL. The developed assay displayed acceptable precision estimated by repeated analysis of the quality control samples and the clinical samples analyzed by this assay showed a good correlation with that commercial kit (r = 0.998).     

A simplified radioimmunoassay for triiodothyronine (T3) using pre-incubated labelled antigen and antibody

We discribe the development of a simplified radioimmunoassay for triiodothyronine (T₃) using pre-incubated labelled T₃ and antibody. The assay is carried out by adding 50 l of standard or sample to 0.4 ml of pre-incubated reagent dispensed in assay tubes. The reaction is allowed to proceed for about four hours and the antigen-antibody complex precipitated by the addition of 1 cm³ of 22% polyethylene glycol solution. Due to the high dissociation constant of T₃-antibody complex at 37° C (2.83·10^–4 S^–1), the labelled antigen-antibody complex dissociates and thereby the unlabelled antigen binds with the antibody. With a four hour incubation the sensitivity of this assay is comparable to an assay done by the equilibrium method using the same antibody. Sixty serum samples were analyzed using this method and compared with the equilibrium assay (Y=0.94x+0.046 ng/cm³, r=0.98).     

Coupling of effector molecules to solid supports

In view of the limited stability of the isourea bond, formed in ligand coupling to CNBractivated polysaccharides, an alternative to this current activation method has been developed. 2,4,6-Trifluoro-5-chloropyrimidine (FCP), known as a reactive group in reactive dyes, was used to activate Sepharose. Under appropriate conditions a thermally stable product with unimpaired beaded structure was obtained, which was reactive toward amines and mercaptans. Coupling with hexamethylenediamine, aniline, and ethanethiol, respectively, yielded an incorporation of 0.2-2.7, 0.9-1.7, and 1.1 mmol ligand/g dry agarose. The stability of immobilized ligands based on FCP-Sepharose between pH 4 and 8 was about 200 times higher as compared to products originating from CNBr-Sepharose; ligand leakage was only 0.5 x 10^p-3%/h. The possibility of obtaining a high degree of substitution is a further advantage of the FCP activation. In addition, the FCP-activated Sepharose can be stored in the wet state at 4°C without substantial decrease in coupling capacity. The FCP analogs 2,4,5,6-tetrachloro- and 2,4,5,6-tetrafluoropyrimidine, and other polymers (cellulose, Sephadex, aminomethylpolystyrene) appeared to be applicable also.     

固相合成法制备纳米碳酸锶微球

以六水合氯化锶(SrCl2·6H2 O)和碳酸氢铵(NH4 HCO3)为原料,十二烷基苯磺酸钠(SDBS)为表面活性剂,采用阴离子表面活性剂辅助固相球磨法制备纳米碳酸锶微球.样品采用扫描电镜(SEM)、透射电镜(TEM)、能谱仪(EDX)、X-射线衍射仪、红外光谱FTIR、热重仪和差示扫描仪联用(TG—DSC)等手段表...     

Evolution of solid phase homochirality for a proteinogenic amino acid

The inexorable evolution of solid-phase single chirality is demonstrated for the first time for a proteinogenic amino acid. Enantioenrichment is observed both under attrition-enhanced conditions and without the aid of particle grinding. Differences in the form of the conversion profiles for the process under the two sets of conditions provide suggestions concerning the mechanism of the transformation.     

Purification of 3-indolylacetic acid by solid phase extraction

This paper deals with the use of solid-phase extraction (SPE) for the extraction and purification of 3-indolylacetic acid (IAA), the main and most important representative of the plant growth regulators auxins. The procedure is based on the use of C18-SPE columns in two steps. In the first one, raw extract in methanol:water (4:1) is applied on the column in order to remove neutral ballast. In the second step the eluate is diluted with water to a final methanol concentration of 20% (to decrease the elution strength of the solvent) and acidified with formic acid to a final concentration of 1% (to convert the IAA from the anionic to the neutral form). Neutral IAA is then retained on the second SPE column, eluted by (acidified) methanol, and subjected to final analysis. Scintillation counting of tritium labeled IAA standard was used for the investigation and optimization of the SPE procedure. Gas chromatography with mass spectrometric detection was suggested as a convenient method for routine determination of IAA after its methylation with diazomethane. Overall recovery of the procedure was estimated as 89-94% and a physiological level of IAA equal to 0.48 nmol/g (84 ng/g) fresh weight was found using an optimized SPE-GC-MS method.     

A traceless solid phase synthesis of thiomorpholin-3-ones

A novel synthesis of thiomorpholin-3-ones using a traceless solid phase approach is described, in which many kinds of thiomorpholin-3-ones were efficiently obtained in high purity based on an intramolecular alkylation of sulfides followed by an elimination of desired thiomorpholin-3-ones from the generated sulfonium salts.     

Solid phase radioimmunoassay for testosterone in human serum using antibodies coupled to magnetizable cellulose

Summary A simple user-friendly, solid phase radioimmunoassay for testosterone in human serum based on magnetic particles is described. IgG fractions precipitated from polyclonal testosterone antiserum were coupled to magnetizable cellulose particles using carbonyl diimidazole. The prepared antibody solid phase was stable for one year when stored at 4 °C. The optimized assay involves the incubation of 50 ml of testosterone standards (0.3-10 ng/ml), 100 ml of magnetizable cellulose particle coupled antibody suspension and 100 ml of ¹²⁵I-testosterone derivative for 4 hours at 37 °C. At the end of the incubation, the tubes were placed on a magnetic rack for 10 minutes after the addition of wash buffer and decanted. The sensitivity of the assay is 0.2 ng/ml. The intra-assay variation was <15% throughout the assay range. The recovery varied from 88 to 115%. On dilution of samples having high levels of testosterone, linearity ranged between 87 and 125%. Correlation coefficient of 0.978 was obtained when the developed solid phase assay was compared to the earlier established liquid phase assay.     

Gas chromatography-mass spectrometry determination of 3-mercaptohexan-1-ol and 3-mercaptohexyl acetate in wine: a comparison of headspace solid phase microextraction and solid phase extraction methods

A gas chromatography-mass spectrometry method was established using headspace solid phase microextraction (HS-SPME) as the sampling procedure to analyse 3-mercaptohexan-1-ol (3-MH) and 3-mercaptohexyl acetate (3-MHA), two molecules with a tropical fruit aroma, in wine at trace level. This method offers important advantages, as it neither requires time-consuming sample preparation nor uses dangerous organic compounds, thus making control of wine aroma easier and suitable for routine analysis. As a comparison, a solid phase extraction (SPE) method, already described elsewhere for aroma analysis, was applied to quantify these analytes, extending this exhaustive enrichment to two important thiols. Detection limits for both the approaches were close to the sensory threshold value, resulting lower for the HS-SPME procedure and suitable for requirements in the oenological field. The application of the two proposed methods to 52 wines of different varieties gave similar results.     

Site-specific covalent attachment of an engineered Z-domain onto a solid matrix: An efficient platform for 3D IgG immobilization

Immobilized antibodies with oriented and homogeneous patterns are crucial to solid-phase molecular recognition assay. Antibody binding protein-based immobilization can effectively present the desired antibodies. However, steadily installing the stromatoid protein with site-specific attachment manner onto a matrix surface remains to be elucidated. In this study, we present an optimal protocol to tightly attach an immunoglobulin G (IgG)-binding protein (Z-domain) through covalent incorporation of Cys-tag and maleimide group onto polystyrene surface to guarantee site-specific, oriented, and irreversible attachment, resulting in a highly efficient platform for three-dimensional IgG immobilization. The actual IgG-binding characteristic of immobilized Z-Cys was investigated by employing affinity chromatography and size exclusion chromatography. And the efficacy and potential of this platform was demonstrated by applying it to the analysis of interaction between rabbit anti-HRP IgG and its binding partner HRP. The proposed approach may be an attractive strategy to construct high performance antibody arrays and biosensors given that the antibody is compatible with the Z-domain.     

A new solid phase approach for rapid syntheses of oligonucleotides bearing a 3′-terminal phosphate group

Synthesis of pentanucleotide ApTpCpTpTp containing a 3′-terminal phosphomonoester group was accomplished with the help of the new phosphate-solid phase link 2-(4-carboxyphenyl-mercapto)ethanol; a similar molecule, 2-benzylsulfonylethanol, revealed interesting features as a temporary 3′-phosphate protecting group.