Use of non-contact hopping probe ion conductance microscopy to investigate dynamic morphology of live platelets.

Circulating platelets are anucleated and multi-functional cells that participate in hemostasis and arterial thrombosis. Multiple ligands and mechanical forces activate platelets, leading to cytoskeletal rearrangement and dramatic shape-changes. Such dramatic changes in platelets membrane structures are commonly detected by optical and electron microscopy after platelets are fixed. We have recently developed a method to study the membrane morphology of live platelets using Hopping Probe Ion Conductance Microscopy (HPICM). We have successfully used this technology to study the process of platelet microvesiculation upon exposure to selective agonists. Here, we further discussed technical details of using HPICM to study platelet biology and compared results from HPICM to those from conventional atomic force microscopy and scanning electron microscopy. This method offers several advantages over current technologies. First, it monitors morphological changes of platelets in response to agonists in real time. Second, platelets can be repeatedly scanned over time without damages brought by heat and prolong light exposure. Third, there is no direct contact with platelet surface so that there will no or minimal mechanical damages brought by a cantilever of a conventional atomic force microscopy. Finally, it offers the potential to study platelet membrane ion channels, which have been technically challenging up-to-date. Our data show that HPICM has high-resolution in delineating changes of platelet morphology in response to stimulations and could help to unravel the complex role of platelet in thrombus formation.