Schubert Rajib, Strohmeyer Nico, Bharadwaj Mitasha, Ramanathan Subramanian P, Krieg Michael, Friedrichs Jens, Franz Clemens M, Muller Daniel J
Department of Biosystems Science and Engineering, ETH Zurich, Mattenstrasse 26, 4058 Basel, Switzerland.
Department of Molecular and Cellular Physiology, Stanford University, Stanford, CA 94305, USA.
FEBS Lett. 2014 Oct 1;588(19):3639-48. doi: 10.1016/j.febslet.2014.06.012. Epub 2014 Jun 10.
Single-cell force spectroscopy (SCFS) is becoming a widely used method to quantify the adhesion of a living cell to a substrate, another cell or tissue. The high sensitivity of SCFS permits determining the contributions of individual cell adhesion molecules (CAMs) to the adhesion force of an entire cell. However, to prepare adherent cells for SCFS, they must first be detached from tissue-culture flasks or plates. EDTA and trypsin are often applied for this purpose. Because cellular properties can be affected by this treatment, cells need to recover before being further characterized by SCFS. Here we introduce atomic force microscopy (AFM)-based SCFS to measure the mechanical and adhesive properties of HeLa cells and mouse embryonic kidney fibroblasts while they are recovering after detachment from tissue-culture. We find that mechanical and adhesive properties of both cell lines recover quickly (<10 min) after detachment using EDTA, while trypsin-detached fibroblasts require >60 min to fully recover. Our assay introduced to characterize the recovery of mammalian cells after detachment can in future be used to estimate the recovery behavior of other adherent cell types.
单细胞力谱（SCFS）正成为一种广泛应用的方法，用于量化活细胞与底物、另一个细胞或组织之间的黏附力。SCFS的高灵敏度使得能够确定单个细胞黏附分子（CAMs）对整个细胞黏附力的贡献。然而，为了制备用于SCFS的贴壁细胞，必须首先将它们从组织培养瓶或培养板上分离下来。为此，常使用乙二胺四乙酸（EDTA）和胰蛋白酶。由于这种处理会影响细胞特性，细胞在通过SCFS进行进一步表征之前需要恢复。在这里，我们引入基于原子力显微镜（AFM）的SCFS，以测量HeLa细胞和小鼠胚胎肾成纤维细胞在从组织培养物中分离后恢复过程中的力学和黏附特性。我们发现，使用EDTA分离后，两种细胞系的力学和黏附特性在分离后迅速恢复（<10分钟），而用胰蛋白酶分离的成纤维细胞需要>60分钟才能完全恢复。我们引入的用于表征哺乳动物细胞分离后恢复情况的检测方法，未来可用于估计其他贴壁细胞类型的恢复行为。
