在生理血流条件下靶向超声微泡对P-选择素的靶向黏附效能

目的平行板流动腔评价在生理血流条件下携抗小鼠P-选择素单抗靶向超声微泡（MBp）的靶向黏附效能。方法采用＂亲和素-生物素＂桥接法构建MBp;在3种浓度（10、100和1000ng/ml）小鼠P-选择素Fc段（PSFc）包被的平行板流动腔和固定剪切应力下,以及最大包被浓度和不同剪切应力（0.2-1.7dyn/cm^2）下分别检测MBp的每分钟结合数量（结合率）,以抗小鼠P-选择素单抗封闭组和空白组为对照。在3种包被浓度下检测MBp达半数解离的剪切应力。所有分组样本数均为3。结果两对照组均未见有明显的MBp结合。实验组MBp结合率随包被浓度的增高而增加（P〈0.05）,然而与剪切应力呈现双向性（P〈0.05）。MBp达半数解离的剪切应力随包被浓度的增加而增大（P〈0.05）。结论MBp在生理条件下可与PSFc特异有效地结合,体外对靶向超声微泡的靶向黏附效能评价将有助于判定超声分子成像的效果和靶向微泡的在体应用环境条件。

Binding capability of microbubbles targeted to P-selectin under physiologic flow conditions

Objective To assess the binding capability of microbubbles targeted to P-selectin using the parallel plate flow chamber under physiologic flow conditions. Methods Targeted microbubble was designed by conjugating monoclonal antibodies against mouse P-selectin to the lipid shell of the microbubble via an "avidin-biotin" bridge. The binding and retention of targeted microbubbles to P-selectin immobilized on a culture dish were assessed in a parallel-plate flow chamber. Targeted microbubbles drawn through the flow chamber coated with P-selectin (10, 100 and 1000 ng/ml) at a shear stress of 0.3 dyn/cm², and different shear stress (0.2-1.7 dyn/cm²) with 1000 ng/ml P-selectin substrates. Control experiments were performed on plates with no P-selectin and on P-selectin coated plates blocked with excess monoclonal antibodies against P-selectin. The retention of microbubbles was tested by ramping up shear stress at 30 s intervals. Results A marked binding of targeted microbubbles was seen, but not in both control groups. With the increase in the plate surface densities of P-selectin, the increase in the binding rate and the shear stress of half detachment of targeted microbubbles were gradually increased (P<0.05). While the shear stress was under 0.5 dyn/cm², the binding rate of targeted microbubbles in the 1000 ng/ml P-selectin substrates was increased with the increase of shear stress (P<0.05). Above that, the binding rate was decreased (P<0.05). Conclusion Binding and retention of the targeted microbubbles to P-selectin is specific and effective under physiologic flow conditions. In vitro evaluation of binding capability of targeted microbubbles could be useful for predicting the effects of molecular ultrasound imaging and the using circumstance of microbubbles in vivo.