Determination of four quinolones in plasm and urine by using mixed mode solid phase extraction and capillary electrophoresis / 国际检验医学杂志

Objective To develop a new method for the rapid determination of lomefloxacin,gatifloxacin,ciprofloxacin and oflox-acin in plasm and urine by solid phase extraction(SPE)and capillary electrophoresis.Methods The capillary was fused silica capil-lary with id/od of 75/365 μm and effective/total length of 40/47 cm.The running buffer was 40 mmol/L borate buffer at pH 9.0. Separation voltage was 13 kV.Temperature was 20 ℃.Detection wave-length was set at 280 nm.The sample was analyzed after the pretreatment of SPE.Results The analysis of lomefloxacin,gatifloxacin,ciprofloxacin and ofloxacin was completed in 6 minutes with satisfied accuracy and precision.Good linearity was found within the range of 1-40 μg/mL,and the r was 0.998 7,0.997 6, 0.998 3 and 0.994 2 respectively.The recoveries of four quinolones in plasm and urine ranged from 80.1% to 107.6%,and the rel-ative standard deviations(RSD)ranged from 2.1% to 6.2%.Conclusion This method is fast,simple,precise and it might be feasi-ble for the determination of lomefloxacin,gatifloxacin,ciprofloxacin and ofloxacin in plasm and urine samples.

Establishment of the methodology for quantifying lentiviral vector transcriptional read-through rate / 生物工程学报

As an effective vehicle for bio-research and for gene therapy, Lentiviral Vector (LV) has been drawn large attention in recent years. However, transcriptional read-through limits its application. In order to understand the extend of LV read-through in chromosome, a reliable method to assess transcriptional read-through rate is needed. Here, we report the method as follows: 293T cells were transfected with the lentiviral transfer vectors which borne with two LTRs at its two ends in order to mimic the state of "proviral vectors" in chromosome. Using the primers specific for 3'U5 and 3'U3, read-through and total transcripts were reverse transcribed, respectively. These two cDNAs were quantified by realtime PCR using the primers and probe specific for 5'end of 3'U3. Read-through rate was then calculated by the division of the two. Meanwhile, read-through product of green fluorescence protein was also analyzed by Fluorescence Activated Cell Sorter. They both reciprocally proved the principal and confirmed that self-inactivated LV appeared higher read-through rate than the wild type one. The method described in this article, therefore, provides a useful technique to study how to reduce read-through rate, and improve the bio-safety of LV.

Progress in improvement of lentiviral vectors' transcriptional read-through / 生物工程学报

Four out of 10 patients of X-linked severe combined immunodeficiency (X-SCID) were finally developed leukemia after receiving the treatment of gene therapy delivered by gamma-retroviral vectors. This is due to the vector integrated to the proximity of lmo2 etc proto-oncogene promoters, leading to the activation of onco-gene expression, which raises the concern of the bio-safety of gene therapy vectors. Lentiviral vectors, especially self-inactivating lentiviral vectors, are considered to be much safer than gamma-retroviral vectors. However self-inactivating lentiviral vectors also have encountered with some unsafe factors and one of them is the problem of transcriptional "read-through" . During the past years, achievements have been made to reduce lentiviral vector transcriptional read-through, which are reviewed herein.