Résumé
@#The assay method of GLP-1 receptor binding affinity for a long-acting hypoglycemic peptide—PEgylated Exendin-4 analogue(PE)was optimized and established based on the luciferase reporter gene approach. CHO-GLP-1R-CRE-Luc+ cells were previously constructed in our lab followed by the verification of methodology. This assay method showed good specificity and robustness as well as high accuracy and precision when PE was incubated with the cell for 4 h, the luminescent substrate reacted with cell lysates for 15 min and the concentration for PE ranged 5. 7×10-3-1. 5×103 nmol/L, on which condition this developed method is in accordance with General Principles of Analytical Method Validation Techniques for Biological Products Quality Control. This study also lays the foundations for rapid evaluation and screening of GLP-1 receptor agonist drugs.
Résumé
Objective To evaluate the targeting and photodynamic activity of photosensitizer Ⅰ (PSⅠ)for laryngocarcinoma Hep-2 cells.Methods The photostability of PSⅠ was measured by bleaching assay.The cellular uptake of PSⅠ by Hep-2 cells were evaluated by fluorescence spectroscopy,the effects of PSⅠconcentration,irradiation dose and etc on the viability of Hep-2 cells were investigated by MTT assay.Results After irradiation of 50 min,the OD value of PS Ⅰ decreased 11％,which indicated that the stability of PSⅠ can meet the requirement of PDT.The cellular uptake of PS Ⅰ by Hep-2 cells increased with the concentration of PSⅠ and can be obviously inhibited by the presence of excessive free folic acid (P ＜ 0.01).The results of MTT assays demonstrated that the viability of Hep-2 showed negative correlation with the PSⅠ concentration and irradiation dose.The viability of Hep-2 was only 34 ％ after PDT with 14 μ mol/L of the PS Ⅰ and 18 J/cm2 of irradiation.However,the viability of Hep-2 was still 100 ％ without irradiation,even though the concentration of PS Ⅰ was up to 1 10 μmol/L.The PDT carried out after 24 h of PSⅠ administration can efficiently inhibited the growth of Hep-2 with the survival rate of 32 ％.Conclusion The PS Ⅰ posses satisfactory photostability and lower dark cytotoxicity,and displays significant targeting and photocytotoxicity for Hep-2 cells.