目的依据血浆中存在游离DNA的理论,采用磁珠作为固相吸附载体并使用特定设计的试剂体系及提取流程,建立一种简便、高效提取尿液样本中游离甲基化DNA的方法,评价其作为用于尿液样本的甲基化基因检测技术的可行性。方法使用磁珠法提取40...目的依据血浆中存在游离DNA的理论,采用磁珠作为固相吸附载体并使用特定设计的试剂体系及提取流程,建立一种简便、高效提取尿液样本中游离甲基化DNA的方法,评价其作为用于尿液样本的甲基化基因检测技术的可行性。方法使用磁珠法提取40例成人尿液中游离甲基化DNA,进行甲基化修饰后,紫外分光光度计测定DNA的浓度和纯度。结果结果显示,提取50 m L尿液可得61~200 ng/μL的甲基化DNA,OD260/280为1.8±0.05。使用甲基化阳性对照DNA引物进行PCR及电泳后可见目的条带,说明其纯度可以满足后续甲基化基因检测和PCR等操作要求。结论尿液中确实存在游离甲基化DNA,同时磁珠法提取尿液游离DNA提取过程简单,且提取纯度高,是一种高效的提取方法。展开更多
Circulating free nucleic acids; cell free DNA and circulating micro-RNA, are found in the plasma of patients with hematologic and solid malignancies at levels higher than that of healthy individuals. In patients with ...Circulating free nucleic acids; cell free DNA and circulating micro-RNA, are found in the plasma of patients with hematologic and solid malignancies at levels higher than that of healthy individuals. In patients with hematologic malignancy cell free DNA reflects the underlying tumor mutational profile, whilst micro-RNAs reflect genetic interference mechanisms within a tumor and potentially the surrounding microenvironment and immune effector cells. These circulating nucleic acids offer a potentially simple, non-invasive, repeatable analysis that can aid in diagnosis, prognosis and therapeutic decisions in cancer treatment.展开更多
We measured the concentrations and distribution of major polyamines(spermine, putrescine and spermidine) in seawater during successive spring algal blooms in an area of frequent harmful blooms in the East China Sea. S...We measured the concentrations and distribution of major polyamines(spermine, putrescine and spermidine) in seawater during successive spring algal blooms in an area of frequent harmful blooms in the East China Sea. Spermine, putrescine, and spermidine concentrations were analyzed by high performance liquid chromatography and ranged from 1–64, 7–81, and 0–19 nmol/L. Spermine was present at the highest concentrations, followed by putrescine and spermidine. In late April, when a diatom bloom dominated by Skeletonema costatum dispersed, polyamine concentrations increased, presumably as a result of diatom decomposition. In early May, when a dinoflagellate bloom dominated by Prorocentrum donghaiense occurred, the polyamine concentration decreased from the level seen in late April. The abundant polyamines that decomposed and were released during the diatom bloom in late April may have promoted the growth of P. donghaiense, resulting in its dominance.展开更多
Lamotrigine (LTG) is a widely used antiepileptic drug (AED) for the treatment of new-onset, as well as refractory epilepsy. Due to the narrow treatment window and large individual variability in the pharmacokinetics a...Lamotrigine (LTG) is a widely used antiepileptic drug (AED) for the treatment of new-onset, as well as refractory epilepsy. Due to the narrow treatment window and large individual variability in the pharmacokinetics and pharmacodynamics of LTG, therapeutic drug monitoring (TDM) is necessary in clinical practice to guide dose adjustments. Individual differences and drug combinations can also affect protein binding rate, which further affects the unbound concentration of LTG. The unbound fraction is more closely related to adverse reactions and therapeutic efficacy than total concentration. Therefore, it may be more meaningful to determine the unbound LTG concentration in plasma than the total concentration.Unbound LTG in plasma was extracted by ultrafiltration. High-performance liquid chromatography (HPLC) was used to measure unbound LTG concentration. This method was validated by studies of its selectivity, linearity, lower limit of quantification (LLOQ), accuracy, precision, recovery, and stability.The method was validated over a linear range of 0.2 to 10.0 μg·mL–1, and its LLOQ was 0.2 μg·mL–1. The method’s relative standard deviations (RSDs) for intra-day and inter-day precision were less than 15%, and its accuracy (RE) was ±4.69%. The recoveries of unbound LTG at three different concentrations satisfied the requirements for the analysis of biological samples, and no significant degradation of LTG was observed under different storage conditions.A simple HPLC method showed good performance when used to measure unbound LTG concentration. This method might be used to study the relationship between unboundLTG concentrations and its effectiveness according to TDM.展开更多
文摘目的依据血浆中存在游离DNA的理论,采用磁珠作为固相吸附载体并使用特定设计的试剂体系及提取流程,建立一种简便、高效提取尿液样本中游离甲基化DNA的方法,评价其作为用于尿液样本的甲基化基因检测技术的可行性。方法使用磁珠法提取40例成人尿液中游离甲基化DNA,进行甲基化修饰后,紫外分光光度计测定DNA的浓度和纯度。结果结果显示,提取50 m L尿液可得61~200 ng/μL的甲基化DNA,OD260/280为1.8±0.05。使用甲基化阳性对照DNA引物进行PCR及电泳后可见目的条带,说明其纯度可以满足后续甲基化基因检测和PCR等操作要求。结论尿液中确实存在游离甲基化DNA,同时磁珠法提取尿液游离DNA提取过程简单,且提取纯度高,是一种高效的提取方法。
文摘Circulating free nucleic acids; cell free DNA and circulating micro-RNA, are found in the plasma of patients with hematologic and solid malignancies at levels higher than that of healthy individuals. In patients with hematologic malignancy cell free DNA reflects the underlying tumor mutational profile, whilst micro-RNAs reflect genetic interference mechanisms within a tumor and potentially the surrounding microenvironment and immune effector cells. These circulating nucleic acids offer a potentially simple, non-invasive, repeatable analysis that can aid in diagnosis, prognosis and therapeutic decisions in cancer treatment.
基金Supported by the National Basic Research Program of China(973 Program)(No.2010CB428701)the National Natural Science Foundation of China(Nos.40976047,42176118)
文摘We measured the concentrations and distribution of major polyamines(spermine, putrescine and spermidine) in seawater during successive spring algal blooms in an area of frequent harmful blooms in the East China Sea. Spermine, putrescine, and spermidine concentrations were analyzed by high performance liquid chromatography and ranged from 1–64, 7–81, and 0–19 nmol/L. Spermine was present at the highest concentrations, followed by putrescine and spermidine. In late April, when a diatom bloom dominated by Skeletonema costatum dispersed, polyamine concentrations increased, presumably as a result of diatom decomposition. In early May, when a dinoflagellate bloom dominated by Prorocentrum donghaiense occurred, the polyamine concentration decreased from the level seen in late April. The abundant polyamines that decomposed and were released during the diatom bloom in late April may have promoted the growth of P. donghaiense, resulting in its dominance.
基金Liaoning Province Natural Science Foundation of China(Grant No.2013021079)
文摘Lamotrigine (LTG) is a widely used antiepileptic drug (AED) for the treatment of new-onset, as well as refractory epilepsy. Due to the narrow treatment window and large individual variability in the pharmacokinetics and pharmacodynamics of LTG, therapeutic drug monitoring (TDM) is necessary in clinical practice to guide dose adjustments. Individual differences and drug combinations can also affect protein binding rate, which further affects the unbound concentration of LTG. The unbound fraction is more closely related to adverse reactions and therapeutic efficacy than total concentration. Therefore, it may be more meaningful to determine the unbound LTG concentration in plasma than the total concentration.Unbound LTG in plasma was extracted by ultrafiltration. High-performance liquid chromatography (HPLC) was used to measure unbound LTG concentration. This method was validated by studies of its selectivity, linearity, lower limit of quantification (LLOQ), accuracy, precision, recovery, and stability.The method was validated over a linear range of 0.2 to 10.0 μg·mL–1, and its LLOQ was 0.2 μg·mL–1. The method’s relative standard deviations (RSDs) for intra-day and inter-day precision were less than 15%, and its accuracy (RE) was ±4.69%. The recoveries of unbound LTG at three different concentrations satisfied the requirements for the analysis of biological samples, and no significant degradation of LTG was observed under different storage conditions.A simple HPLC method showed good performance when used to measure unbound LTG concentration. This method might be used to study the relationship between unboundLTG concentrations and its effectiveness according to TDM.
