Kang Hyen-Wook, Otani Naoya, Hiroshi Muramatsu, Chang Sang-Mok, Kim Jong Min
Department of Chemical Engineering, Dong-A University, Busan, 604-714, Korea.
Department of Bioscience and Biotechnology, Tokyo University of Technology, Tokyo, 192-0982, Japan.
J Nanosci Nanotechnol. 2018 Aug 1;18(8):5777-5784. doi: 10.1166/jnn.2018.15466.
A real-time quartz crystal microbalance (QCM) cell activity monitoring system coupled with micro CCD cameras was developed to investigate the cultured cell activity, which could measure the viscoelastic characteristics of the cell with the QCM and observe the cell morphology changes with CCD camera simultaneously. Both the viscoelastic characteristics and the shape of the cultured cell are important factors to estimate the cell activity and the cell adhesion. The extracellular matrix (ECM) on the surface of the QCM is essential to culture the cell stably in the QCM monitoring system. To find the ECM optimization condition, the adhesive strength of cultured cells on the ECM modified glass surface was measured by using rotating water stream and CCD camera. After culturing HepG2 cells for 24 hours on the ECM modified glass plates, the glass plates were dipped in the PBS solution and rotated with 1,000, 1,300, and 1,500 rpm for 30 seconds. The adhesiveness of ECMs was investigated by calculating the remained cells after rotating. Four types of ECM, such as amino group, carboxyl group, collagen monomer, and collagen polymer, were used and tested. The current paper improves the sensing system of previous report so that measurements of four ECMs can be simultaneously conducted under the same conditions in order to enhance reliability. A collagen polymer exposed ECM was the most stable on an adhesiveness point of view, but not suitable for the QCM cell activity monitoring due to the decrease of the QCM sensitivity. The sensitivity of the QCM cell activity monitoring system using collagen monomer as ECM is about 2.6 times better than that using collagen polymer. A collagen monomer exposed ECM was more stable than amino group and carboxyl group exposed ECMs based on an adhesiveness point of view. Therefore, a collagen monomer exposed ECM was the most stable and suitable for the QCM cell activity monitoring system among the four ECMs. The changes of the resonance frequency and the resonance resistance of the ECM film with the cultured cells were investigated and compared the results of CCD camera images. From these results, we showed the QCM cell activity monitoring system coupled with the micro CCD camera could be applied to the evaluation of the cell activities.
开发了一种结合微型电荷耦合器件（CCD）相机的实时石英晶体微天平（QCM）细胞活性监测系统，用于研究培养细胞的活性，该系统能够通过QCM测量细胞的粘弹性特征，并同时利用CCD相机观察细胞形态变化。培养细胞的粘弹性特征和形状都是评估细胞活性和细胞黏附的重要因素。QCM表面的细胞外基质（ECM）对于在QCM监测系统中稳定培养细胞至关重要。为了找到ECM的优化条件，使用旋转水流和CCD相机测量了培养细胞在ECM修饰玻璃表面的黏附强度。在ECM修饰的玻璃板上培养HepG2细胞24小时后，将玻璃板浸入磷酸盐缓冲盐水（PBS）溶液中，并分别以1000、1300和1500转/分钟的速度旋转30秒。通过计算旋转后剩余的细胞数量来研究ECM的黏附性。使用了四种类型的ECM，即氨基、羧基、胶原单体和胶原聚合物，并进行了测试。本文改进了先前报告中的传感系统，以便能够在相同条件下同时对四种ECM进行测量，以提高可靠性。从黏附性角度来看，暴露有胶原聚合物的ECM最稳定，但由于QCM灵敏度降低，不适合用于QCM细胞活性监测。使用胶原单体作为ECM的QCM细胞活性监测系统的灵敏度比使用胶原聚合物的系统高出约2.6倍。从黏附性角度来看，暴露有胶原单体的ECM比暴露有氨基和羧基的ECM更稳定。因此，在这四种ECM中，暴露有胶原单体的ECM最稳定，最适合用于QCM细胞活性监测系统。研究了带有培养细胞的ECM膜的共振频率和共振电阻的变化，并比较了CCD相机图像的结果。从这些结果可以看出，结合微型CCD相机的QCM细胞活性监测系统可应用于细胞活性评估。
