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Since its inception, anti-platelet antibodies have been an important tool for studying the interaction of platelets with blood components and blood vessels. At the same time, anti-platelet antibodies also play an important role in the detection and diagnosis of hemorrhagic and thrombotic diseases, and become a kind of powerful anti-thrombotic drugs. With the further understanding of the role of platelets in physiological hemostasis and pathological thrombosis, anti-platelet and anti-thrombotic antibodies also seek a balance between better anti-thrombotic effects and less bleeding side effects.
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Objective To evaluate and report the performance of PL-11 platelet analyzer.Methods Intravenous blood samples anticoagulated with EDTA-K2 and sodium citrate were tested by the PL-11 platelet analyzer to evaluate the intra-assay and inter-assay coefficient of variation (CV),carry-over rate,accuracy,linearity of the PL-11 platelet analyzer.Platelet aggregation rate of sodium citrate-anticoagulated fasting venous blood collected from 30 physical examinees in outpatient department of The First Affiliated Hospital of Soochow University between February and July,2012 was detected by the PL-11 platelet analyzer and MPG-3E multifunctional double channel blood coagulation analyzer,respectively.The correlation was detected between the PL-11 platelet analyzer and MPG-3E multifunctional double channel blood coagulation analyzer.Results All the parameters were conformed to the standard of Clinical Laboratory Improvement Amendment 88.Both of the intra-assay and inter-assay CV values were less than 5％ ; carry-over rate was less than 1％ ; the accuracy and the linearity was excellent correlated to the result of photoelectric turbidimetry (R2 =0.9439).Conclusions PL-11 platelet analyzer can directly use whole blood dynamics to analyze platelet aggregation process and quantitatively analyze the various components of blood cells (including platelets and red blood cells),the result reports are accurate and reliable.
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Objective:To label monoclonal anti-GPⅢa antibody (SZ21) with 99 Tcm and evaluate the value of 99 Tcm-SZ-21 for detecting thrombi in vivo.Methods:SZ-21 was modified with 2-iminotholane and labeled with 99 Tcm-GH and radiochemical purity was determined by ITLC-SG.99Tcm-SZ-21 was injected via ear edge vein in 5 rabbits in which thrombi were induced in the right femoral arteries.As control,99 Tcm-GH was administered in 1 rabbit.Static images were acquired and irregularly shaped ROIs were drawn on the images to calculate the ratios of T/M and T/B.Vein blood was drawn at 2 min,5 min,10 min,30 min and 1 h-3 h after injection in 2 rabbits so as to observe the blood clearance of 99 Tcm-SZ-21.Rabbits were sacrificed after 3 h of imaging.The vessels including clots were harvested and imaged.Cardiac muscle,liver,spleen,lung,kidney,etc.were excised and weighed.Radioactivity counts were measured to calculate % ID/g.Results:The radiochemical purity of 99 Tcm-SZ21 was beyond 90% and stable in vitro.Thrombi could be visualized at 30 min after injection,and at 2 h image of thrombi was clearly visualized,T/M (2.55?0.72) and T/B (1.94?0.51) ratios were high.In vitro imaging showed that T/B was 4.43?1.5.Conclusion:99Tcm-SZ-21 could be a potential agent for imaging diagnosis of thrombotic disease.
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AIM: To further investagate the mechanism of thrombus formation and develop a new remedy of anti-thrombus formation. METHODS: The amplified DNA fragment of vWF-A1 domain was inserted into expression vector with 6?his taq (pQE-31), the recombinant expression vect or was transformed into E coli (strain M15) and induced by IPTG. The recombinant fragment, comprising residues 449-728 of mature vWF subunit, designate rvWF-A1. It was purified by Ni-NTA agarose column and renatured by Tris buffer containin g GSH and GSSG. FACS and platelet aggregometer were employed to analyse the rvWF -A1 function of binding to platelet glycoprotein Ib and inhibiting ristocetin-in duced platelet aggregation. RESULTS: The rvWF-A1 was expressed successfully in E coli, comin g up to 30% of total bacterial protein. Its purify was over 95% through Ni-NTA a garose. It was identified to have ability to bind to GPIb, its biologic activity to inhibit ristocetin-induced platelet aggregation was observed, and the inhibi tive rate was 84 7%. CONCLUSION: The above results indicated that high-level expressi on of rvWF-A1 was successfully achieved in E coli and rvWF-A1 may be an effectiv e antithromotic agent in preventing thrombus formation.
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AIM: To investigate the expression of tissue factor(TF) induced by oxidized high density lipoprotein(oxHDL) in human umbilical vein cell line,ECV304,and the related mechanisms.METHODS: Four main groups were designed: the negative,the positive(ECV304 with histamine),the HDL group and the oxHDL group.Quantitative real-time polymerase chain reaction(RQ-PCR) and Western blotting were used to detect the expression level of TF.The specific inhibitors of MAPKs,SP600125(c-jun terminal NH2 kinase,JNK),SB203580(p38 MAP kinase,p38 MAPK),PD98059(extracellular signal-regulated kinase,ERK1/2) were used to investigate the underlying mechanisms.RESULTS: The TF expression in normal ECV304 cell line was not detected.Histamine administration resulted in a significant expression of TF in ECV304 cell line,with strongest effect after 1 h co-incubation at concentration of 1?10-5 mol/L histamine(about 4.8-fold higher expression of TF compared with that of 1?10-9 mol/L histamine).Expression level of TF was detected after stimulated with oxHDL in dose-and time-dependent manners.The highest expression of TF mRNA was found at 20 mg/L oxHDL and 6 h co-incubation,with 1.8-fold and 5.3-fold increase in TF expression,respectively,compared with that at 10 mg/L oxHDL and 2 h co-incubation.20 mg/L oxHDL also caused an apparent augmentation of TF protein expression,about 1.5-fold higher compared with that stimulated by 40 mg/L oxHDL.HDL co-incubation did not cause a detectable expression of TF protein.The mRNA levels of TF in ECV304 cell line induced by oxHDL were decreased by 95.0%,81.0%,87.0%,respectively(all P