Search results for: Rabbit Anti-Phospho-NFKB1(Ser927) Polyclonal Antibody, Unconjugated
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A novel multiplex method for the simultaneous detection and relative quantitation of pollen allergens.
Standardization of pollen protein extracts is essential in order to ensure efficiency and safety in allergy diagnosis and immunotherapy. In this paper, we have optimized a multiplex Western blotting method for the simultaneous detection of four olive pollen allergens (Ole e 1, Ole e 2, Ole e 5, and Ole e 9) on a single blot using a monoclonal antibody from mouse and three polyclonal antibodies raised in rabbit. We utilized unconjugated Fab antibody fragments for blocking rabbit primary antibodies, and fluorescence-based detection. These changes allowed an accurate and reliable comparative quantitation of these allergens among pollen-protein samples from six olive cultivars. In addition, we also tested the IgE-binding capacity of these pollen extracts by reprobing the same blot with a pool of sera from eight patients allergic to olive and detection with enzyme conjugated antibodies. A noticeable variability regarding allergen content and IgE-reactivity was found among the olive cultivars analyzed. Moreover, we could easily confirm the identity of some of the IgE-binding proteins by simply overlapping both fluorescence and chemiluminescence images. This method is versatile since it can be applied to other allergogenic plant species and extended to other allergens.Sonia Morales, Antonio Jesús Castro, Jose Carlos Jimenez-Lopez, Fernando Florido, María Isabel Rodríguez-García, Juan de Dios Alché
2076 related Products with: A novel multiplex method for the simultaneous detection and relative quantitation of pollen allergens.
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#17462880 2007/03/23
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A novel radioimmunoassay of 16alpha-hydroxy-dehydroepiandrosterone and its physiological levels.
16alpha-Hydroxy-dehydroepiandrosterone (16alpha-OH-DHEA) belongs to the products of extensive DHEA metabolism in mammalian tissues. It is a precursor of 16alpha-hydroxylated estrogens, increased levels of which are associated with autoimmune disorders. A highly specific radioimmunoassay of unconjugated 16alpha-OH-DHEA was developed and evaluated. Polyclonal rabbit antisera were raised against 3beta,16alpha-dihydroxy-17,19-dione-19-O-(carboxymethyloxime) and 3beta,16alpha-dihydroxy-7,17-dione-7-O-(carboxymethyloxime) BSA conjugates. Two methods were used for preparation of the conjugates. Homologous radioiodinated derivatives with tyrosine methyl ester were prepared as tracers. While antisera to 7-CMO cross-reacted with DHEA as much as by 58%, the cross-reaction of the chosen antiserum prepared via 19-oxogroup by micellar conjugation technique with 16beta-OH-DHEA was only 0.13% and with all other structurally related steroids, including DHEA were lower than 0.01%. The detection limit was 0.017 pmol (5.7 pg)/tube, the average intra- and inter-assay coefficients of variation were 8.2 and 11.4%, respectively. Mean recovery of serum spiked with 16alpha-OH-DHEA varied between 80 and 110%, the results were independent on sample dilution. 16alpha-OH-DHEA concentrations in 18 randomly selected sera, including 6 samples from patients with thyroid cancer were compared with results obtained by earlier GC-MS method. Physiological levels of 16alpha-OH-DHEA in 316 sera (184 females and 132 males) analyzed so far varied between 0.0 and 1.86 nmol/l.Ludmila Zamrazilová, Hana Kazihnitková, Oldrich Lapcík, Martin Hill, Richard Hampl
2854 related Products with: A novel radioimmunoassay of 16alpha-hydroxy-dehydroepiandrosterone and its physiological levels.
5 G 25 MG1 mg1 mg 5 G10 mg0.1 ml100 mg1 ml500 mg10 mg200ul
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#17298836 2006/12/20
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A novel radioimmunoassay of 7-oxo-DHEA and its physiological levels.
A novel radioimmunoassay (RIA) of unconjugated 7-oxo-dehydroepiandrosterone (7-oxo-DHEA) in human serum was developed for the first time. This steroid is an intermediate in the biosynthesis of immunomodulatory 7-hydroxylated DHEA metabolites, and has been shown to possess thermogenic properties. The method employs polyclonal rabbit antiserum to (19E)-3beta-hydroxy-7,17,19-trione-19-O-(carboxymethyloxime):BSA conjugate and a homologous radioiodinated derivative with tyrosine methyl ester. The cross reactivity of the antiserum with structurally closest 7-hydroxyepimers of DHEA was lower than 1.7%, with DHEA 0.4%, with all other related steroid less than 0.4%. The method includes ether extraction of serum (0.5 ml), followed by RIA. Its detection limit was 0.06 pmol (18 pg)/tube, the average intra- and inter-assay coefficients of variation were 4.1% and 8.3%, respectively. Mean recovery of serum spiked with 7-oxo-DHEA varied between 78.8% and 112%. Its levels in three serum pools were compared with a low-resolution gas chromatography-mass spectrometry method with satisfactory results. The method has been used for determination of 7-oxo-DHEA in serum samples of 215 subjects (91 males and 124 females) without overt endocrine disorders, aged 5-71 years. The over-all mean+/-S.D. was 0.280+/-0.227, the median 0.239 nmol/l. No significant sex differences were recorded. The only group which differed significantly from all other ones were males below 10 years, significantly lower values than in other age groups were found also in the first two age groups of females.H Kazihnitková, L Zamrazilová, M Hill, O Lapcík, V Pouzar, R Hampl
1427 related Products with: A novel radioimmunoassay of 7-oxo-DHEA and its physiological levels.
200ug1,000 tests50 ug 1 mg1000 TESTS/0.65ml1 ml25 mg100ul1000 tests2.5 mg1,000 tests
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Enantioselective immunorecognition of protein modification with optically active ibuprofen using polyclonal antibody.
Formation of covalently bound protein adducts with 2-arylpropionic acids (2-APAs) has been proposed as a possible explanation for hypersensitivity and toxic responses to chiral carboxylic acid drugs. To identify the cellular proteins chemically modified with optically active (S)-ibuprofen, we generate polyclonal antibodies by immunizing rabbits with immunogen coupled to bovine serum albumin (BSA) via the spacer of 4-aminobutyric acid. The resulting antibodies largely cross-reacted with N-alpha-(t-butoxycarbonyl)--(S)-ibuprofenyl lysine as well as with the conjuguated (S)-ibuprofen with glycine and taurine and unconjugated (S)-ibuprofen, enabling enantioselective detection of (S)-ibuprofen residues anchored on ovalbumin molecules, introduced by the reaction of the ibuprofen p-nitrophenyl ester. Furthermore, immunoblotting with an antibody allows the enantioselective detection of (S)-ibuprofen-introduced glutathione-S-transferase (GST). These results indicate that the developed method will be useful for monitoring the generation and localization of protein covalently bound with (S)-ibuprofen, which may be the cause of ibuprofen-induced toxicity.Hiromi Ito, Shunji Ishiwata, Takeshi Kosaka, Rika Nakashima, Harunori Takeshita, Sakiko Negoro, Masako Maeda, Shigeo Ikegawa
1210 related Products with: Enantioselective immunorecognition of protein modification with optically active ibuprofen using polyclonal antibody.
100ug Lyophilized100ug Lyophilized100ug Lyophilized100ug Lyophilized100ug Lyophilized100ug Lyophilized100ug Lyophilized100ug Lyophilized100ug Lyophilized100ug Lyophilized100ug Lyophilized100ug Lyophilized
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#9190879 //
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Immunochemical assay for recognition of 2-S-glutathionyl acetate, a glutathione conjugate derived from 1,1-dichloroethylene-epoxide.
Cytotoxicities induced by 1,1-dichloroethylene (DCE) are ascribed to cytochrome P450-dependent metabolism to an epoxide. Conjugation of the DCE-epoxide with glutathione (GSH) results in the formation of the conjugates 2-S-glutathionyl acetate (GTA) and 2-(S-glutathionyl) acetyl glutathione (GAG); GAG undergoes hydrolysis to form GTA, and thus GTA is a major metabolite of DCE metabolism. Our objective is to develop an antiserum against the chemically synthesized GTA, and for immunization, we have used a hapten that consists of GTA conjugated to bovine serum albumin (BSA) as the carrier protein and glutaraldehyde (GLUT) as a chemical cross-linker. The antisera were raised in rabbits and were characterized by using the following synthesized structural analogs: GTA, glycine-GLUT-BSA (GLY-GLUT-BSA), GTA-GLUT-ovalbumin (GTA-GLUT-OVB), GTA-1-ethyl-3-(3-dimethylaminopropyl) carbodiimide-BSA (GTA-EDC-BSA), TRIS-GLUT-BSA, glutathione-GLUT-BSA (GSH-GLUT-BSA). The enzyme-linked immunosorbent assay (ELISA) and slot immunoblotting were used to characterize the specificity of the antisera. Noncompetitive ELISA experiments showed that the reaction of the antiserum with the antigen was concentration-dependent. In the competitive ELISA, GTA-GLUT-BSA inhibited binding efficiently; in contrast, the unconjugated GTA did not inhibit binding to the antigen. Competitive studies with the other analogs indicated low or minimal reactivities with the antibodies, which were blocked by incubation with GLY-GLUT-BSA. However, there was residual reactivity with the antigen that was not competitively inhibited by either the GTA-EDC-BSA or the GSH-GLUT-BSA conjugates. Slot-blotting experiments confirmed the findings of the ELISA studies and revealed high specificity of the antiserum to detect the hapten. These results demonstrated the successful development of polyclonal antibodies to detect GTA and hence DCE-epoxide.P G Forkert, K S Collins, T F Dowsley, G M Ross
1345 related Products with: Immunochemical assay for recognition of 2-S-glutathionyl acetate, a glutathione conjugate derived from 1,1-dichloroethylene-epoxide.
1 kit1 L200 assays100Tests250200 assays480 tests100 tests50 assays100 assays100 assays
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#9071741 //
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A novel radioimmunoassay for daidzein.
A radioimmunoassay for daidzein was established, based on polyclonal antibodies against daidzein-4'-O-(carboxymethyl)ether-BSA. The sensitivity of the assay was 0.4 pg/tube; the intra- and interassay coefficients of variation varied from 4.1 to 11.5% and from 5.6 to 21.7%, respectively, depending upon the method (direct or extraction) and concentration of daidzein in the sample. The cross reactivities with other chemically related compounds, with the exception of 4'-derivatives of daidzein, were 2.4% for dihydrodaidzein, 1.3% for genistein, 1.5% for biochanin A, and 1.6% for equol, respectively. The method was used for measurement of daidzein levels in 105 normal human subjects and in three volunteers after consumption of a meal prepared from 125 g of cooked whole soybeans. The daidzein values obtained following diethyl ether extraction of human sera was only 8% of that obtained by direct radioimmunoassay. We suggest that this difference is caused by cross-reacting daidzein 4'-glucuronides and -sulfates present in serum. Using ether extraction, the basal serum levels of free daidzein were 0.11 ng/mL (0.43 nmol/L), with 14 subjects showing no detectable levels. Levels were detectable in all subjects with the direct assay with a mean value of 7.1 ng/mL (28.0 nmol/L). Peak levels were reached 4 hours after ingestion of the soybeans. The levels were 10.3 +/- 3.6 ng/mL (40.4 +/- 14.3 nmol/L) for free daidzein and 129.4 +/- 36.1 ng/mL (509 +/- 142 nmol/L) for total immunoreactive compounds. After 24 hours, the levels were still clearly distinguishable from the basal levels; the concentrations were 0.43 +/- 0.15 ng/mL (1.69 +/- 0.59 nmol/L) and 24.36 +/- 6.07 ng/mL (95.9 +/- 23.9) for free and total immunoreactive material, respectively. It is concluded that this is the first immunoassay for a phytoestrogen in human biological fluids, and for the first time serial assays of unconjugated daidzein in plasma have been possible.O Lapcík, R Hampl, N al-Maharik, A Salakka, K Wähälä, H Adlercreutz
1039 related Products with: A novel radioimmunoassay for daidzein.
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#9116591 //
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An antiserum to idazoxan recognizes an immunoreactive substance in human serum and cerebral spinal fluid which is not agmatine.
A polyclonal antibody was generated in rabbits to an idazoxan-albumin antigen. The anti-idazoxan antiserum had high affinity for unconjugated 3H-idazoxan (Kd of 19.8 nM) in a radio-immunoassay (RIA). Of various drugs and native molecules only idazoxan potently (Ki of 24 nM) inhibited 3H-idazoxan binding to the anti-idazoxan antibody. A few drugs weakly inhibited 3H-idazoxan binding (IC50 > 605 microM) with rank order of UK 14304 > guanabenz > cirazoline > amiloride > naphazoline. Neither agmatine, an endogenous clonidine displacing substance (CDS), catecholamines or imidazoles inhibited the binding of 3H-idazoxan to the anti-idazoxan antibody. The anti-idazoxan RIA was 4-6 fold more sensitive than an antibody to para-amino clonidine. The CDS detected by ligand displacement from bovine brain dose-dependently inhibited 3H-idazoxan binding. This immunoreactive (ir-) CDS activity was present in human (0.9-4.1 U/ml) and rat sera (1-2 U/ml) and in the cerebro-spinal fluid of eight patients with serious disease of the central nervous system, but not in controls. We conclude: (1) an anti-idazoxan RIA is a sensitive, selective and clinically applicable RIA for measuring ir-CDS; (2) ir-CDS is not agmatine; (3) CDS represents a family of endogenous ligands for imidazoline receptors including ir-CDS and agmatine.H Wang, S Regunathan, D McGowan, S Bramwell, D J Reis
1580 related Products with: An antiserum to idazoxan recognizes an immunoreactive substance in human serum and cerebral spinal fluid which is not agmatine.
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#8288443 //
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Polyclonal anti-idiotypes induce specific anti-saxitoxin antibody responses.
Polyclonal BALB/C mouse and New Zealand White rabbit anti-idiotypic antibodies were raised by immunization with a protein G-purified burro anti-saxitoxin IgG antibody preparation. Following absorption of non-anti-idiotype reactivity, murine and rabbit IgG were purified by protein A chromatography and used to immunize BALB/C mice for the induction of anti-saxitoxin antibody responses. Unconjugated BALB/C anti-idiotypes did not induce significant anti-saxitoxin reactivity in BALB/C mice, even after repeated immunizations. However, BALB/C mice immunized with purified BALB/C anti-idiotypes conjugated to keyhole limpet hemocyanin, or with purified, unconjugated rabbit anti-idiotypes, as aluminum hydroxide precipitates, induced significant and specific anti-saxitoxin immune responses. Saxitoxin, a sodium channel blocker, can protect cells treated with veratridine and ouabain, whose respective actions are to open sodium channels and to block the activity of Na/K-ATPase. The anti-idiotype-induced anti-saxitoxin antibodies inhibited saxitoxin from protecting against cell death induced by veratridine and ouabain treatment. These and other published experimental results strengthen the concept of anti-idiotype-based vaccines in eliciting protective immunity against a variety of low molecular weight, nonproteinaceous biological and chemical toxins, whose extreme toxicity does not allow their use as safe immunogens.T C Chanh, J F Hewetson