In mammalian skeletal muscle there are four carboxylases involved in several biochemical processes like gluconeogenesis, tricarboxylic acid cycle anaplerosis, metabolism of fatty acids and metabolism of various amino ...In mammalian skeletal muscle there are four carboxylases involved in several biochemical processes like gluconeogenesis, tricarboxylic acid cycle anaplerosis, metabolism of fatty acids and metabolism of various amino acids. It has been shown that biotin deficiency reduces body weight at the expense of muscular mass. When necessary, the liver uses skeletal muscle protein to provide glucose and amino acids to organs in need of such compounds. In this paper we analyzed carboxylase specific activities in hind limb skeletal muscle of 3 weeks old BALB/c male mice, at 0, 1, 4, 7, and 14 days of a specific diet with different biotin concentrations. Biotin was used at 0.0, 1.8 or 98.2 mg per kg of food;and was referred to as biotin deficient, sufficient and supplemented, respectively. Water and food supply and consumption by the three groups of mice were the same. Therefore, the observed effects were directly related to biotin ingestion. The body weight of biotin supplemented mice was the same as the body weight of mice in the biotin sufficient group, while biotin deficiency caused body weight reduction after 7 days of biotin depletion. We found that the total protein concentration in the vastus lateralis muscle is associated with the biotin content in the diet. After 7 days, the muscle total protein content was lower in mice of the biotin deficient group while it was higher in the mice from the biotin supplemented group展开更多
Results from 4 switchback field trials involving 608 cows were combined to assess the effects of a protected B vitamin blend (BVB) vs 10 mg of unprotected biotin upon milk yield (kg), fat %, protein %, fat yield (kg) ...Results from 4 switchback field trials involving 608 cows were combined to assess the effects of a protected B vitamin blend (BVB) vs 10 mg of unprotected biotin upon milk yield (kg), fat %, protein %, fat yield (kg) and protein yield (kg) in primiparous and multiparous cows. Trials consisted of 3 DHIA periods executed in the order control-test-control. Cows from 45 to 300 days in milk (DIM) at the start of the experiment that were available for all 3 periods were included in the analysis. No diet changes other than the substitution of 3 grams/cow/day of the BVB for 10 mg of biotin during the test period occurred. Results from the two control periods were compared to results obtained during the test period by individual cow using a paired T test. Results for all cows showed that the provision of the BVB resulted in increased (P < 0.05) milk, fat percentage (%), protein %, fat yield and protein yield. Analysis by age revealed that milk production and milk protein yield were only improved in mature cows. Milk production had a negative effect upon the magnitude of the increase in milk components. The change in milk yield was greatest in early lactation and declined with DIM. Protein % and fat % increased with DIM in mature cows, but not in first lactation cows. Differences in fat yields between test and control feeding periods did not change with DIM, but the improvement in protein yield in mature cows declined with DIM. These results indicate that the BVB provided economically important advantages throughout lactation beyond those witnessed with biotin, but expected results would vary with cow age and stage of lactation.展开更多
Background: Peripheral neuropathy is a commonly encountered troublesome condition which is often disabling & worsens when left untreated. Traditional neuropathic pain medications primarily provide symptomatic reli...Background: Peripheral neuropathy is a commonly encountered troublesome condition which is often disabling & worsens when left untreated. Traditional neuropathic pain medications primarily provide symptomatic relief;however, the pathogenesis of nerve damage remains unresolved. Extensive literature survey reveals that patients with peripheral neuropathy experience significant benefits with the use of B-vitamins like methylcobalamin (B12), folic acid (B9), biotin (B7), benfotiamine (B1) and pyridoxine (B6). The other well documented antineuropathic agents include alpha lipoic acid, glutathione, omega fatty acids, myoinositol, certain trace elements, etc. Materials and Methods: A multicentre, prospective, open-label, non-comparative clinical study was carried out in 497 patients with peripheral neuropathy. A fixed dose combination of methylcobalamin, alpha lipoic acid (ALA), folic acid, biotin, benfotiamine & vitamin B6 capsule was orally administered once daily for 12 weeks. Results: Treatment led to significant reduction from baseline score in various neuropathy symptoms from the 4th week itself. After 12 weeks of treatment, the mean pain score declined by 78.0%, numbness by 92.1% and muscle weakness by 96.9%. Also, there was 96.0% & 99.2% reduction in tingling & burning sensation respectively. No serious adverse events were reported.Conclusion: The current study confirms that fixed dose combination of methylcobalamin, ALA, folic acid, biotin, benfotiamine & vitamin B6 is effective & well tolerated in the management of peripheral neuropathy.展开更多
Two biotinylated derivatives of the fungal metabolite galiellalactone (1) were synthesized in order to facilitate the investigation of the molecular mechanism of action of the galiellalactonoids. Galiellalactone is a ...Two biotinylated derivatives of the fungal metabolite galiellalactone (1) were synthesized in order to facilitate the investigation of the molecular mechanism of action of the galiellalactonoids. Galiellalactone is a STAT3-signaling inhibitor that inhibits growth in vitro as well as in vivo of prostate cancer cells expressing activated STAT3. To provide a suitable point of attachment for biotin, the 8-hydroxymethyl derivative (3) and its 7-phenyl analogue 4 were synthesized by a modified tandem Pd-catalysed carbonylation and intramolecular vinyl allene Diels-Alder procedure previously developed. The two primary alcohols obtained, 3 and 4, were coupled to biotin as the 6-aminohexanoic acid amide, activated as the acid chloride, yielding the derivatives 5 and 6.展开更多
Succinic acid is valued as a potential starting point for the production of chemicals of the C4 family or in the prepara-tion of biodegradable polymers. For sustainable development in this era of petroleum shortage, p...Succinic acid is valued as a potential starting point for the production of chemicals of the C4 family or in the prepara-tion of biodegradable polymers. For sustainable development in this era of petroleum shortage, production of succinic acid by microbial fermentation of renewable feedstock has attracted great interest. In this study, pretreatment with sulfuric acid and biotin supplementation were used to enhance succinic acid production by Actinobacillus succinogenes 130Z from sake lees, a byproduct of Japanese rice wine. Pretreatment with sulfuric acid resulted in little change of glucose, total nitrogen and succinic acid content in the sake lees hydrolysate but had a positive effect on succinic acid fermentation, which caused a 25.0% increase in succinic acid yield in batch fermentation. Biotin supplementation was used to further enhance the fermentability of sake lees hydrolysate. As a result, a 30 h batch fermentation of 0.5% sulfuric acid pretreated sake lees hydrolysate with 0.2 mg/L biotin gave a succinic acid yield of 0.59 g/g from 61.6 g/L of glucose, with a productivity of 1.21 g/(L?h). A 22.9% increase in succinic acid yield and a 101.7% increase in succinic acid productivity were obtained compared with untreated sake lees hydrolysate.展开更多
The rupture force of the streptavidin-biotin complex was investigated using atomic force microscopy (AFM). The most frequently observed rupture force (MFOF), which is essential for the evaluation of the potential land...The rupture force of the streptavidin-biotin complex was investigated using atomic force microscopy (AFM). The most frequently observed rupture force (MFOF), which is essential for the evaluation of the potential landscape, was evaluated by processing 22,500 force curves using two methods. One method is a conventional method, which is usually built in commercial AFM systems, i.e., difference between the baseline value and the minimum force value in the force curve. The other is a detection of rupture events based on a fuzzy logic algorithm to detect the rupture event from analyzing the shape of the force curves. Our statistical analysis revealed that the conventional method exhibited a significant artifact, which is the increase in the population of small forces comparable to thermal noise of cantilevers, resulting in a smaller MFOF. Based on this finding, we discuss the choice of a method and its effecton the illustrated potential landscapes of ligand-receptor complexes.展开更多
AIM: To evaluate the multi-step pretargeting radioimmunoimaging (RII) and radioimmunotherapy (RIT) in nude mice bearing human colon carcinoma with avidin-biotin system labeled with 153Sm.METHODS: Two- and three-step s...AIM: To evaluate the multi-step pretargeting radioimmunoimaging (RII) and radioimmunotherapy (RIT) in nude mice bearing human colon carcinoma with avidin-biotin system labeled with 153Sm.METHODS: Two- and three-step strategies for avidinbiotin system pretargeting techniques were established.In a three-step procedure, human colon carcinoma bearing nude mice were first injected with biotinylated monoclonal antibody (McAb-Bt) followed by cold avidin (Av) 48 h later and then 153Sm-DB2 24 h thereafter;whereas the twostep procedure consisted of injection of 153Sm-SA 48 h after pretargeting with biotinylated anti-CEA monoclonal antibody (CEA McAb-Bt). SPECT imaging and biodistribution were performed at 4, 24, 48, or 72 h after injection of 153Sm-labeled compounds. Five groups of nude mice subcutaneously grafted with human colon carcinoma were treated 3 d after grafting. One group received the injection with 100 μg CEA McAb-Bt followed by cold avidin (80 μg)after 2 d and 11.1 MBq 153Sm-DB2 after 1d. Four control groups were treated respectively with 11.1 MBq 153SmCEA McAb, 11.1 MBq 153Sm-nmIgG, 11.1 MBq 153Sm-DB2,100 Μl normal saline. Toxicity was evaluated by changes of leukocyte count, and the efficacy by variation in tumor volume. Histological analyses of tumors were performed.RESULTS: The three-step procedure allowed faster blood clearance and yielded higher tumor blood ratios (5.76 at4 h and 12.94 at 24 h) of the 153Sm-DB2. The tumor was clearly visualized at 4 h in y-imaging after the injection of 153Sm-DB2, while a significant accumulation of 153Sm-SA in the tumor was observed only 24 h after the injection and tumor blood ratios at 4 and 24 h were 1.00 and 2.03,respectively, in the two-step procedure. Pretargeting RIT and 153Sm-CEA McAb had a strong tumor-inhibiting effect.The tumor inhibitory rate was 80.67% and 78.44%,respectively, five weeks after therapy. Histopathological evidence also indicated radioactive damage in tumor tissues as necrosis of tumor cells, while in the other organs such as liver and kidney no radioactive damage was observed. Leukocyte counts showed significant decrease after treatment in groups of 153Sm-CEA Mc Ab and 153SmnmIgG.CONCLUSION: The two kinds of pretargeting strategies can elevate the target-to-nontarget ratio, decrease the blood background and shorten the imaging time compared to 153Sm-CEA McAb. Three-step pretargeting RIT is as efficient as 153Sm-CEA Mc Ab, but markedly less toxic. This study provides experimental evidence for the clinical application of pretargeting RII and RIT.展开更多
Objective To report a protocol using biotin-labelled PrP protein in cell free conversion assay instead of isotope. Methods A hamster PrP protein (HaPrP) was expressed in E. coli and purified with HIS-tag affinity chro...Objective To report a protocol using biotin-labelled PrP protein in cell free conversion assay instead of isotope. Methods A hamster PrP protein (HaPrP) was expressed in E. coli and purified with HIS-tag affinity chromatograph. After being labelled with biotin, HaPrP was mixed with PrPSc preparation from scrapie strain 263K. Results Protease-resistant bands were detected after four-day incubation. Conclusion The new conversion model provides a reliable, easily handling, and environment-friendly method for studies of prion and transmissible spongiform encephalopathies.展开更多
Aim To develop a sensitive competitive ELISA for the determination of biotin in transformed yeast culture media.Methods The ELISA plate was firstly coated with Mycoplasma hyopneumoniae, and then successively incubated...Aim To develop a sensitive competitive ELISA for the determination of biotin in transformed yeast culture media.Methods The ELISA plate was firstly coated with Mycoplasma hyopneumoniae, and then successively incubated with rabbit ami-Mycoplasma hyopneumoniae serum and goat anti-rabbit IgG-biotin to form the solid biotin, which competed with the biotin in the solution (standard or sample) for the limited streptavidin-horse radish peroxidase conjugate. The standard calibration curve for biotin analysis was constructed in the range of 50-2000ng·L^-1. Results The detection limit for biotin was found to be 83 ng·L^-1 , which waa about 1000 times lower than the lowest determination concenlration in the reported ELISA for biotin analysis. The relative standard deviations for the spiked samples at biotin concerarations of 200 ng·L^-1, 500 ng·L^-1 , and 1000 ng·L^-1 were 24.87%, 6.15%, and 7.86%, respectively, with the average recovery of 101.13%. The wild yeast and its sixty-three transformed yeast culture media were applied to the developed ELISA for the determination of biotin. It was found that the biotin concentrations in more than 85 % of the tested samples were enhanced with different increase factors after transformation. Conclusion Utilization of Mycoplasma hyopnetunoniae as the coating protein improves the precision and accmacy oftbe ELISA assay, which might be used for the biotin assay in other media.展开更多
A sensitive flow-injection chemiluminescence method was developed for the determination of biotin in the pharmaceutical formulations.The affinity between avidin and biotin was used to adsorb biotin on the polystyrene,...A sensitive flow-injection chemiluminescence method was developed for the determination of biotin in the pharmaceutical formulations.The affinity between avidin and biotin was used to adsorb biotin on the polystyrene,with subsequent quantification of biotin based on its ability to enhance the chemiluminescence(CL) signal generated by the redox reaction of potassium permanganate-luminol-CdTe nanoparticles CL system.The investigations prove that apart from 3-aminophthalate,the CdTe quantum dots(QDs) play both catalytic and emitter roles.Under optimum conditions,the linear range for the determination of biotin was 0.01―25 ng/mL with a detection limit of 7.3×10-3 ng/mL(S/N=3).The relative standard deviation of 5 ng/L biotin was 2.06%(n=7).The proposed method was used to determine the biotin concentration in the pharmaceutical formulations and the recovery was between 96.4% and 104%.The proposed method is simple,convenient,rapid and sensitive.展开更多
The specific binding of receptor to ligand covalently attached to surface with different surface densities was studied using streptavidin-biotin model pair. Biotinylated substrates with different spacer thicknesses as...The specific binding of receptor to ligand covalently attached to surface with different surface densities was studied using streptavidin-biotin model pair. Biotinylated substrates with different spacer thicknesses as formed through a simple reaction between amine immobilized surfaces and N-hydroxysucciimide groups at the end of biotin modifi ed PEG in anhydrous organic solutions("grafting to" technique). The amount of the specifi cally adsorbed protein was measured as a function of spacer thickness between hard surface and biotin moieties. It has been shown that the amount of specifically adsorbed streptavidin decreases with the increase spacer thickness and the protein adsorbs onto the functionalized surfaces in a single molecular manner. It provides an interesting model system for studying single molecular interactions.展开更多
Activity-based Ubiquitin probes(Ub-ABPs)carrying a reporter group have emerged as effective tools for the investigation of deubiquitinating enzymes(DUBs),such as studying the molecular mechanism of DUBs,profiling new ...Activity-based Ubiquitin probes(Ub-ABPs)carrying a reporter group have emerged as effective tools for the investigation of deubiquitinating enzymes(DUBs),such as studying the molecular mechanism of DUBs,profiling new DUBs.But so far,the synthesis of commonly used biotin-bearing Ub-ABPs is a technical challenge.Here,we report a one-pot semi-synthetic strategy for the acquiring of Ub-ABPs carrying a biotin tag through sequential enzymatic ligation,N-S acyl transfer and aminolysis reaction without any purification steps.These probes enable to capture the different family of DUBs for enrichment and immunoblotting using the attached biotin tag.展开更多
基金supported by a grant from the Direccion General de Asuntos del Personal Academico(DGAPA),Universidad Nacional Autonoma de Mexico(PAPIIT IN215311).MA P MdeO was fellow of DGAPA.
文摘In mammalian skeletal muscle there are four carboxylases involved in several biochemical processes like gluconeogenesis, tricarboxylic acid cycle anaplerosis, metabolism of fatty acids and metabolism of various amino acids. It has been shown that biotin deficiency reduces body weight at the expense of muscular mass. When necessary, the liver uses skeletal muscle protein to provide glucose and amino acids to organs in need of such compounds. In this paper we analyzed carboxylase specific activities in hind limb skeletal muscle of 3 weeks old BALB/c male mice, at 0, 1, 4, 7, and 14 days of a specific diet with different biotin concentrations. Biotin was used at 0.0, 1.8 or 98.2 mg per kg of food;and was referred to as biotin deficient, sufficient and supplemented, respectively. Water and food supply and consumption by the three groups of mice were the same. Therefore, the observed effects were directly related to biotin ingestion. The body weight of biotin supplemented mice was the same as the body weight of mice in the biotin sufficient group, while biotin deficiency caused body weight reduction after 7 days of biotin depletion. We found that the total protein concentration in the vastus lateralis muscle is associated with the biotin content in the diet. After 7 days, the muscle total protein content was lower in mice of the biotin deficient group while it was higher in the mice from the biotin supplemented group
文摘Results from 4 switchback field trials involving 608 cows were combined to assess the effects of a protected B vitamin blend (BVB) vs 10 mg of unprotected biotin upon milk yield (kg), fat %, protein %, fat yield (kg) and protein yield (kg) in primiparous and multiparous cows. Trials consisted of 3 DHIA periods executed in the order control-test-control. Cows from 45 to 300 days in milk (DIM) at the start of the experiment that were available for all 3 periods were included in the analysis. No diet changes other than the substitution of 3 grams/cow/day of the BVB for 10 mg of biotin during the test period occurred. Results from the two control periods were compared to results obtained during the test period by individual cow using a paired T test. Results for all cows showed that the provision of the BVB resulted in increased (P < 0.05) milk, fat percentage (%), protein %, fat yield and protein yield. Analysis by age revealed that milk production and milk protein yield were only improved in mature cows. Milk production had a negative effect upon the magnitude of the increase in milk components. The change in milk yield was greatest in early lactation and declined with DIM. Protein % and fat % increased with DIM in mature cows, but not in first lactation cows. Differences in fat yields between test and control feeding periods did not change with DIM, but the improvement in protein yield in mature cows declined with DIM. These results indicate that the BVB provided economically important advantages throughout lactation beyond those witnessed with biotin, but expected results would vary with cow age and stage of lactation.
文摘Background: Peripheral neuropathy is a commonly encountered troublesome condition which is often disabling & worsens when left untreated. Traditional neuropathic pain medications primarily provide symptomatic relief;however, the pathogenesis of nerve damage remains unresolved. Extensive literature survey reveals that patients with peripheral neuropathy experience significant benefits with the use of B-vitamins like methylcobalamin (B12), folic acid (B9), biotin (B7), benfotiamine (B1) and pyridoxine (B6). The other well documented antineuropathic agents include alpha lipoic acid, glutathione, omega fatty acids, myoinositol, certain trace elements, etc. Materials and Methods: A multicentre, prospective, open-label, non-comparative clinical study was carried out in 497 patients with peripheral neuropathy. A fixed dose combination of methylcobalamin, alpha lipoic acid (ALA), folic acid, biotin, benfotiamine & vitamin B6 capsule was orally administered once daily for 12 weeks. Results: Treatment led to significant reduction from baseline score in various neuropathy symptoms from the 4th week itself. After 12 weeks of treatment, the mean pain score declined by 78.0%, numbness by 92.1% and muscle weakness by 96.9%. Also, there was 96.0% & 99.2% reduction in tingling & burning sensation respectively. No serious adverse events were reported.Conclusion: The current study confirms that fixed dose combination of methylcobalamin, ALA, folic acid, biotin, benfotiamine & vitamin B6 is effective & well tolerated in the management of peripheral neuropathy.
文摘Two biotinylated derivatives of the fungal metabolite galiellalactone (1) were synthesized in order to facilitate the investigation of the molecular mechanism of action of the galiellalactonoids. Galiellalactone is a STAT3-signaling inhibitor that inhibits growth in vitro as well as in vivo of prostate cancer cells expressing activated STAT3. To provide a suitable point of attachment for biotin, the 8-hydroxymethyl derivative (3) and its 7-phenyl analogue 4 were synthesized by a modified tandem Pd-catalysed carbonylation and intramolecular vinyl allene Diels-Alder procedure previously developed. The two primary alcohols obtained, 3 and 4, were coupled to biotin as the 6-aminohexanoic acid amide, activated as the acid chloride, yielding the derivatives 5 and 6.
文摘Succinic acid is valued as a potential starting point for the production of chemicals of the C4 family or in the prepara-tion of biodegradable polymers. For sustainable development in this era of petroleum shortage, production of succinic acid by microbial fermentation of renewable feedstock has attracted great interest. In this study, pretreatment with sulfuric acid and biotin supplementation were used to enhance succinic acid production by Actinobacillus succinogenes 130Z from sake lees, a byproduct of Japanese rice wine. Pretreatment with sulfuric acid resulted in little change of glucose, total nitrogen and succinic acid content in the sake lees hydrolysate but had a positive effect on succinic acid fermentation, which caused a 25.0% increase in succinic acid yield in batch fermentation. Biotin supplementation was used to further enhance the fermentability of sake lees hydrolysate. As a result, a 30 h batch fermentation of 0.5% sulfuric acid pretreated sake lees hydrolysate with 0.2 mg/L biotin gave a succinic acid yield of 0.59 g/g from 61.6 g/L of glucose, with a productivity of 1.21 g/(L?h). A 22.9% increase in succinic acid yield and a 101.7% increase in succinic acid productivity were obtained compared with untreated sake lees hydrolysate.
文摘The rupture force of the streptavidin-biotin complex was investigated using atomic force microscopy (AFM). The most frequently observed rupture force (MFOF), which is essential for the evaluation of the potential landscape, was evaluated by processing 22,500 force curves using two methods. One method is a conventional method, which is usually built in commercial AFM systems, i.e., difference between the baseline value and the minimum force value in the force curve. The other is a detection of rupture events based on a fuzzy logic algorithm to detect the rupture event from analyzing the shape of the force curves. Our statistical analysis revealed that the conventional method exhibited a significant artifact, which is the increase in the population of small forces comparable to thermal noise of cantilevers, resulting in a smaller MFOF. Based on this finding, we discuss the choice of a method and its effecton the illustrated potential landscapes of ligand-receptor complexes.
文摘AIM: To evaluate the multi-step pretargeting radioimmunoimaging (RII) and radioimmunotherapy (RIT) in nude mice bearing human colon carcinoma with avidin-biotin system labeled with 153Sm.METHODS: Two- and three-step strategies for avidinbiotin system pretargeting techniques were established.In a three-step procedure, human colon carcinoma bearing nude mice were first injected with biotinylated monoclonal antibody (McAb-Bt) followed by cold avidin (Av) 48 h later and then 153Sm-DB2 24 h thereafter;whereas the twostep procedure consisted of injection of 153Sm-SA 48 h after pretargeting with biotinylated anti-CEA monoclonal antibody (CEA McAb-Bt). SPECT imaging and biodistribution were performed at 4, 24, 48, or 72 h after injection of 153Sm-labeled compounds. Five groups of nude mice subcutaneously grafted with human colon carcinoma were treated 3 d after grafting. One group received the injection with 100 μg CEA McAb-Bt followed by cold avidin (80 μg)after 2 d and 11.1 MBq 153Sm-DB2 after 1d. Four control groups were treated respectively with 11.1 MBq 153SmCEA McAb, 11.1 MBq 153Sm-nmIgG, 11.1 MBq 153Sm-DB2,100 Μl normal saline. Toxicity was evaluated by changes of leukocyte count, and the efficacy by variation in tumor volume. Histological analyses of tumors were performed.RESULTS: The three-step procedure allowed faster blood clearance and yielded higher tumor blood ratios (5.76 at4 h and 12.94 at 24 h) of the 153Sm-DB2. The tumor was clearly visualized at 4 h in y-imaging after the injection of 153Sm-DB2, while a significant accumulation of 153Sm-SA in the tumor was observed only 24 h after the injection and tumor blood ratios at 4 and 24 h were 1.00 and 2.03,respectively, in the two-step procedure. Pretargeting RIT and 153Sm-CEA McAb had a strong tumor-inhibiting effect.The tumor inhibitory rate was 80.67% and 78.44%,respectively, five weeks after therapy. Histopathological evidence also indicated radioactive damage in tumor tissues as necrosis of tumor cells, while in the other organs such as liver and kidney no radioactive damage was observed. Leukocyte counts showed significant decrease after treatment in groups of 153Sm-CEA Mc Ab and 153SmnmIgG.CONCLUSION: The two kinds of pretargeting strategies can elevate the target-to-nontarget ratio, decrease the blood background and shorten the imaging time compared to 153Sm-CEA McAb. Three-step pretargeting RIT is as efficient as 153Sm-CEA Mc Ab, but markedly less toxic. This study provides experimental evidence for the clinical application of pretargeting RII and RIT.
基金This work was supported by National Natural Science Foundation of China 30070038 and 30130070, National High-Tech Research and Development Program of China (863 Project) 2001AA215391, and EU Project QLRT 2000 01441.
文摘Objective To report a protocol using biotin-labelled PrP protein in cell free conversion assay instead of isotope. Methods A hamster PrP protein (HaPrP) was expressed in E. coli and purified with HIS-tag affinity chromatograph. After being labelled with biotin, HaPrP was mixed with PrPSc preparation from scrapie strain 263K. Results Protease-resistant bands were detected after four-day incubation. Conclusion The new conversion model provides a reliable, easily handling, and environment-friendly method for studies of prion and transmissible spongiform encephalopathies.
文摘Aim To develop a sensitive competitive ELISA for the determination of biotin in transformed yeast culture media.Methods The ELISA plate was firstly coated with Mycoplasma hyopneumoniae, and then successively incubated with rabbit ami-Mycoplasma hyopneumoniae serum and goat anti-rabbit IgG-biotin to form the solid biotin, which competed with the biotin in the solution (standard or sample) for the limited streptavidin-horse radish peroxidase conjugate. The standard calibration curve for biotin analysis was constructed in the range of 50-2000ng·L^-1. Results The detection limit for biotin was found to be 83 ng·L^-1 , which waa about 1000 times lower than the lowest determination concenlration in the reported ELISA for biotin analysis. The relative standard deviations for the spiked samples at biotin concerarations of 200 ng·L^-1, 500 ng·L^-1 , and 1000 ng·L^-1 were 24.87%, 6.15%, and 7.86%, respectively, with the average recovery of 101.13%. The wild yeast and its sixty-three transformed yeast culture media were applied to the developed ELISA for the determination of biotin. It was found that the biotin concentrations in more than 85 % of the tested samples were enhanced with different increase factors after transformation. Conclusion Utilization of Mycoplasma hyopnetunoniae as the coating protein improves the precision and accmacy oftbe ELISA assay, which might be used for the biotin assay in other media.
基金Supported by the National Natural Science Foundation of China(No.21075050)the Science and Technology Development Project of Jilin Province,China(No.20110334)
文摘A sensitive flow-injection chemiluminescence method was developed for the determination of biotin in the pharmaceutical formulations.The affinity between avidin and biotin was used to adsorb biotin on the polystyrene,with subsequent quantification of biotin based on its ability to enhance the chemiluminescence(CL) signal generated by the redox reaction of potassium permanganate-luminol-CdTe nanoparticles CL system.The investigations prove that apart from 3-aminophthalate,the CdTe quantum dots(QDs) play both catalytic and emitter roles.Under optimum conditions,the linear range for the determination of biotin was 0.01―25 ng/mL with a detection limit of 7.3×10-3 ng/mL(S/N=3).The relative standard deviation of 5 ng/L biotin was 2.06%(n=7).The proposed method was used to determine the biotin concentration in the pharmaceutical formulations and the recovery was between 96.4% and 104%.The proposed method is simple,convenient,rapid and sensitive.
文摘The specific binding of receptor to ligand covalently attached to surface with different surface densities was studied using streptavidin-biotin model pair. Biotinylated substrates with different spacer thicknesses as formed through a simple reaction between amine immobilized surfaces and N-hydroxysucciimide groups at the end of biotin modifi ed PEG in anhydrous organic solutions("grafting to" technique). The amount of the specifi cally adsorbed protein was measured as a function of spacer thickness between hard surface and biotin moieties. It has been shown that the amount of specifically adsorbed streptavidin decreases with the increase spacer thickness and the protein adsorbs onto the functionalized surfaces in a single molecular manner. It provides an interesting model system for studying single molecular interactions.
基金supported by the National Key R&D Program of China(No.2017YFA0505400)the National Natural Science Foundation of China(Nos.21877024,21972214 and 22277020)。
文摘Activity-based Ubiquitin probes(Ub-ABPs)carrying a reporter group have emerged as effective tools for the investigation of deubiquitinating enzymes(DUBs),such as studying the molecular mechanism of DUBs,profiling new DUBs.But so far,the synthesis of commonly used biotin-bearing Ub-ABPs is a technical challenge.Here,we report a one-pot semi-synthetic strategy for the acquiring of Ub-ABPs carrying a biotin tag through sequential enzymatic ligation,N-S acyl transfer and aminolysis reaction without any purification steps.These probes enable to capture the different family of DUBs for enrichment and immunoblotting using the attached biotin tag.
