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Objective:To validate an enzyme linked immunosorbent assay (ELISA) method for the quantification of rhCNB in long-tailed macaque sera.Methods: The linear,sensitivity,accuracy,precision and recovery were determined using ELISA.Results:The present ELISA method had high linearity within 0.195 ng/ml-12.5 ng/ml,the working curve of rhCNB was Y=15.1X-0.26, R2=0.996 8 , the method showed good sensitivity of 0.195 ng/ml, the accuracy were in the range of 91.9%-108.8%, and the Coefficient of variation ( CV) for inter-assay were 3.55%,1.39%and 4.71%,the intra-assay were 1.59%,3.2%and 3.8%,all less than 10%, the recoveries were in the range of 88.5% -108.3%, <110% .Thus the method was coincidence with requirement.Conclusion:Double antibody sandwich ELISA assay of rhCNB in long-tailed macaque sera has good sensitivity ,accuracy, precision and recovery and it can be used to measure rhCNB concentration in biological samples .
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OBJECTIVE:To establish a method for determining recombinant human calmodulin B subunit(rhCNB)in rat plas-ma,and study its pharmacokinetics characteristics. METHODS:ELISA double-antibody sandwich method was adopted. 1 μg/ml rhCNB monoclonal antibody mAb was wrapped,added to the to-be-test sample,rhCNB polyclonal antibody pAb(dilution ratio of 1∶5 000)and HRP-labeled conjugate of anti-IgG(dilution ratio of 1∶10 000)were added. Using tetramethylbenzidine for develop-ing,microplate reader was conducted in wavelength of 450 nm to determine the absorbance value(OD value)and plasma concen-tration of 6 rats after 2,15,30,60,120,240,480,720 min of iv 2.5 mg/kg rhCNB,and the pharmacokinetic parameters were calculated by BAPP 3.0 software. RESULTS:The linear range of rhCNB were 0.195-12.5 ng/ml(r2=0.995 0),lower limit of quan-titation was 0.195 ng/ml,accuracy were 97.300%-103.622%(RSD<7.5%,n=6);RSDs of within-batch,inter-batch,freezing and thawing 3 times were no higher than 8.5%(n=6,18,15). rhCNB pharmacokinetics characteristics in rat fitted to two-com-partment model,AUC0-720 min was 173.038 mg·min/L and t1/2 was 94.62 min. CONCLUSIONS:The established method has high specificity and sensitivity,good accuracy and precision,which can be used for rhCNB quantitative detection and pharmacokinetics study in biological samples.
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Objective:To investigate the antitumor activity of the compound HS-4 and the action mechanism.Methods:MTT method was used to testin vitroantitumor activity of the compound HS-4. Orthotopic xenotransplantation tumor model of liver cancer was established in nude mice, and,in vivoantitumor activity of compound HS-4 was tested with a small animal in-vivo imaging system. Sequencing of small RNA library and RNA library was performed in HS-4 treated tumor cell group and control group to investigate the anti-cancer mechanism of HS-4 at level of functional genomics, using high-throughput sequencing technology. Results:HS-4 was found to have relatively highin-vitro antitumor activity against liver cancer cells, gastric cancer cells, renal cancer cells, lung cancer cells, breast cancer cells and colon cancer cells. The IC50 values against SMMC-7721 and Bel-7402 of liver cancer cells were 0.14 and 0.13 nmol/L respectively, while the IC50 values against MGC-803 and SGC-7901 of gastric cancer cells were 0.19 and 0.21 nmol/L, respectively. It was demonstrated that HS- 4 possessed a better therapeutic effect in liver cancer.Conclusions: A new reliable orthotopic xenotransplantation tumor model of liver cancer in nude mice is established. The new compounds HS-4 was found to possess relatively highin vivo andin vitroantitumor activity against liver cancer cells.