Fabrication Of Bi-module Sensing Device And Its Application In Tumor Biology

Cell migration is a physical and chemical multistep cycle including extension of a protrusion, formation of stable attachments near the leading edge of the protrusion,translocation of the cell body forward, and release of adhesions and retraction at the cell rear. Cell migration is one of the key cell functions in physiological and pathological processes, especially in tumor metastasis. In recent years some researches have been done in the field of cell migration, mostly part of which are cell migration tests. However, it is not feasible to monitor the important biochemical molecules produced during cell migrations in situ by conventional cell migration assays.Herein, a bi-module sensing device was fabricated to quantify biochemical molecules released from the cell migration process in situ and explore the interaction between tumor cells and immune cells. The device contains both electrochemical sensing and trans-well cell migration modules. The novel bi-module cell migration platform enables in situ investigation of cell secretion and cell function simultaneously,highlighting its potential for characterizing cell motility through monitoring biochemical molecules production on rare samples and for identifying underlying mechanisms of cell migration. Biochemical molecules detected includes ROS（hydrogen peroxide H₂O₂） and cytokines（interferon gamma IFN-γ）. It was aimed to explore the correlation both biological molecules and tumor cell migration and evaluate antitumor immunity in vitro by the analysis of record data. Bi-module sensing device proves a novel method that can characterize cell motility and real-time detect biochemical molecules. The main parts of thesis is as follows:1. Bi-module sensing device to in situ quantitatively detect hydrogen peroxide released from migrating tumor cells The fully assembled device with a multi-wall carbon nanotube/graphene/Mn O2 nanocomposite functionalized electrode was able to successfully characterize hydrogen peroxide（H₂O₂） production from melanoma A375 cells, larynx carcinoma HEp-2 cells and liver cancer Hep G2 under serum established chemotaxis. The maximum concentration of H₂O₂ produced from A375, HEp-2 and Hep G2 in chemotaxis was（130±1.3） n M,（70±0.7） n M and（63±0.7） n M, respectively. The average migration rate of the A375, HEp-2 and Hep G2 cells were（98±6）%,（38±4）% and（32±3）%,respectively. The paper highlights bi-module device potential for characterizing cell motility through monitoring H₂O₂ production on rare samples and for identifying underlying mechanisms of cell migration.2. Detection of IFN-γ released from tumor antigen stimulated immune cells On the purpose of exploration the interaction between tumor cells and immune cell to evaluate antitumor immunity in vitro, bi-module sensing device was used to detect IFN-γ released from immune cells. The limit of IFN-γ detection is 2.45pg/ml with aptamer sensor. The aptamer sensor sensitively responded to IFN-γ and characterized interferon-γ production from Human acute mononuclear leukemia THP-1cell and macrophages RAW264.7 cell under stimulus of factor LPS and liver cancer Hep G2 cell. Relative to immune cells stimulated with LPS, amount of IFN-γ released from immune cells co-cultured with tumor cells was more. In addition, the bi-module device was used to quantitatively detect the concentration of IFN-γ released from THP-1 cell stimulated during tumor cells migration. The results demonstrate that: as a tumor antigen, tumor cells activate the immune cells（THP-1 and RAW264.7）to release IFN-γ.In summary, a bi-module sensing device was fabricated for in situ characterization of small species released from cells during cell migration. The electrochemical performance of H₂O₂ and IFN-γ detection was well characterized. The real-time electrochemical signal caused by small molecules production during cell migration and tumor cell-immune cell interaction were recorded at any time. It was confirmed that hydrogen peroxide is produced in the process of cell migration. The bi-module cell migration platform enables characterize H₂O₂ production in situ while monitoring cell migration capability with rare samples. Tumor cells and immune cells were co-cultured to detect IFN-γ released from immune cells with bi-model device. It proved that activated immune cells released IFN-γ under the stimulation of tumor antigen.