Objective: To develop a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method with cation exchange-based solid phase extraction (SPE) for determination of tetrodotoxin (TTX) in salted pufferfish. Methods: Evenly crushed salted pufferfish samples were subjected to ultrasound-assisted extraction with 0.5% acetic acid/50% methanol/water. The extract was cleaned with cation exchange-based SPE cartridge and eluted with 0.3% hydrochloric acid and 50% acetonitrile/water. The eluent was neutralized with ammonia and separated with a Waters XBridgeTM BEH Amide column (150 mm×3.0 mm, 1.7 μm), and determined using LC-MS/MS in a multiple reaction monitoring mode. Results: The matrix effects of TTX were 85.7%-92.4%, and the matrix suppression effect was under effective control following clean-up procedures using the optimized SPE method. The TTX showed a good linear relationship at the range of 2.0 to 4 000 μg/kg, with a correlation coefficient (r2) of 0.999 2. The limits of detection and quantitation for TTX in sample matrix were 1.0 μg/kg and 2.0 μg/kg, respectively. The mean spiked-recovery rates were 81.2% to 96.5% at spiked amounts of 2.0, 200 μg/kg and 2 200 μg/kg, with relative standard deviations (RSDs) of 4.3% to 7.5%. The intraday accuracy and precision of TTX were 84.4% to 95.6% and 4.9% to 5.8% in quality control samples, and the interday accuracy and precision of TTX were 86.1% to 94.9% and 5.5% to 8.5% in quality control samples. The detection of TTX was 60.5% in 38 market-sold salted pufferfish products using the established LC-MS/MS method. Conclusion The established LC-MS/MS method is effective for accurate quantitative determination of TTX in salted pufferfish.

BACKGROUND:Side population(SP)cells,with varied contents,are widely distributed in adult tissues,embryos,and certain tumor cells.Haematological tumor is one of the main pathological conditions,which endangers human life.Thus,it is important to recognize SP cells in haematological tumor cell lines.OBJECTIVE:To identify whether hematologic tumor cell lines contain SP cells,and to explore a stable detection and separation methods.METHODS:Cells with the characteristics of stem cells being capable of fluorescent dye Hoechst33342 were isolated by flow cytometry;whether there were SP cells in logarithmic growth period of NB4,Raji,K562/ADM and K562 or not were detected.After sorting K562 subpopulations,the purity of sorted cells was detected.RESULTS AND CONCLUSION:After Hoechst33342 staining,flow cytometry results showed that the SP cells appeared in the haematological tumor NB4,Raji,K562/ADM and K562 cell lines,which accounted for 0.8%,2.7%,1.3% and 2.7%,respectively.These cells could be blocked by Verapamil.The purity was greater after a second detection of SP and Non SP cells in K562 cells.

Objective To set up real-time quantitative RT-PCR technique and measure leukemia fusion gene transcripts in patients with AML of FAB-M_2 subtype,and also to investigate the positive rate in patients and the relationship between the AMLI/ETO mRNA levels and the response rate after chemical therapy.Methods The plasmid containing the AMLI/ETO fusion gene sequences were constructed from myeloid cell lines Kasumi-1 (expressing AML1/ETO)to establish the standard curves,A TaqMan based realtime quantitative RT-PCR was performed to measure aberrant fusion gene transcripts in 45 samples of peripheral blood(PB) or bone marrow (BM) from 25 newly diagnosed patients with AML-M_2.All these 25 patients were diagnosed by the FCM(flow cytometry)and bone marrow molecular cytogenetics,and received the induce remission therapy with MA (Mitoxantrone+Ara-C).Results As a result,the AML1/ETO fusion gene transcripts were detected in 7(28%)out of 25 AML-M_2 patients (the ratios of AML1/ETO/ABL vary from 0.01 to 19.2),in which 5 patients were found t(8;21)(q22;q22).The transcript level of AML1/ETO fusion gene varied from the clinical situation of patients.These 7 patients with AML1/ETO fusion gene got complete remission(CR) after the first MA therapy,and the fusion gene reduced by 3 log in AML1/ETO/ABL.Only 11 patients got CR in 18 patients without AML1/ETO fusion gene.By following up these 7 patients with AML1/ ETO fusion gene kept persistent CR for 6 months.Conclusion It was concluded that real-time quantitative PCR is a reliable,innovative and promising technology with high sensitivity and speciality.It has potential clinical value for diagnosis,tumor typing,treatment selection,measuring the tumor load,monitoring fusion gene expression level and evaluating therapeutic strategy.It is worthy to apply in the clinical practice.

Objective To investigate the effect of fetoscopic photocoagulation of communicating placental vessels in twin-twin transfusion syndrome(TTTS)(selective or non-selective) on the perinatal outcomes.Methods Six cases of TTTS admitted in our department from Dec.2006 to Jun.2008 underwent fetoscopic photocoagulation of communicating vessels.Under direct real-time sonographic guidance,a 3-mm-diameter fetoscope was percutaneously inserted through the maternal abdominal wall into the amniotic cavity of the recipient twin.A combination of ultrasonographic and fetoscopic vision was used to identify the crossing vessels which were systematically coagulated using Nd:YAG laser fiber or bipolar electrocoagulation.Results All the 6 mothers tolerated the procedure without major complications.Two fetal survival rate was 33.33%.Conclusion Fetoscopic photocoagulation of communicating placental vessels in TTTS can effectively improve perinatal outcomes.