Co-Culturing Cancer Cells and Normal Cells in a Biochip under Electrical Stimulation

Kin Fong Lei,Shao-Chieh Hsieh,Rei-Lin Kuo,Ngan-Ming Tsang 한국바이오칩학회 2018 BioChip Journal Vol.12 No.3

The ideal cancer therapeutic strategy is to inhibit the tumor with minimal influence on the normal tissue. Recently, applying an alternating electric field for inhibiting tumor was developed; but, it has not been adopted to be one of the regular therapeutic options. More basic scientific evidence is needed to clarify the efficacy and safety. In the current study, co-culturing cancer cells and normal cells under the electrical stimulation was conducted to provide evidence of this novel cancer therapy. A microfluidic cell culture biochip has been developed and consisted of nine culture chambers incorporating with stimulating electrodes. Cells cultured in the chamber received uniform electric field and cell viability was studied during the culture course. The electric field perturbs cell division and the correlation between cell proliferation rate and inhibition effect was studied among five cell lines, i.e., Huh7, HeLa, TW06, BM1, and HEL299. The results confirmed that cells with higher proliferation rate responded to a higher inhibition. In addition, co-culturing cancer cells and normal cells was conducted to mimic in vivo microenvironment that consists of both cancer and stromal cells. The cancer cells and normal cells were respectively transduced with green fluorescent protein and red fluorescent protein in order to differentiate the cells in a same culture chamber. During the culture course, the electric field was applied to the culture chamber and both cells simultaneously received the field. The results indicated that the growth of the cancer cells were inhibited while the normal cells were maintained. These results provided the evidence of the therapeutic efficacy and safety. Moreover, the microfluidic cell culture biochip could be used for the systematic and precise investigations of the cellular responses under the electrical stimulation.

Development of a 2-Chamber Culture System for Impedimetric Monitoring of Cell-cell Interaction

Kin Fong Lei,Meng-Tsan Tsai,Ming-Hong Zhong,Chia-Hao Huang,Ngan-Ming Tsang,Ming-Yih Lee 한국바이오칩학회 2017 BioChip Journal Vol.11 No.2

In cancer research, study of cell-cell interaction is important to understand tumor initiation, progression, metastasis, and therapeutic resistance. Conventionally, transwell system was adopted and cell proliferation was quantified by end-point bio-assays. The operations are labor-intensive and time-consuming while studying of the dynamic cellular responses of cell-cell interaction. Although impedance measurement was suggested to be a promising technique to monitor cellular responses, electrodes cannot be integrated into the transwell for the measurement purpose. In this work, a 2-chamber culture system incorporated with impedance measurement technique was developed to quantitatively study cell-cell interaction. The chamber was composed of 2 sub-chambers separated with a barrier. By this design, two types of cells could be independently cultured and concurrently monitored under common medium supplied. Cell-cell interaction was demonstrated by aberrant cell proliferation induced by the EGF secreted from the transfected cells cultured on another sub-chamber. Real-time and non-invasive monitoring of cell-cell interaction was successfully demonstrated. This work provides a practical solution for monitoring the dynamic cellular responses of cellcell interaction during the culture course. It is a reliable and convenient platform and facilitate more quantitative assessments in cancer research.

A Reagent-Ready-on-Chip Microfluidic Immunoassay System for Rapid Detection of Influenza A H1N1 and H3N2 Viruses

Kin Fong Lei,Jun-Liang Liu,Chia-Hao Huang,Rei-Lin Kuo,Ngan-Ming Tsang 한국바이오칩학회 2016 BioChip Journal Vol.10 No.1

Development of effective disease screening method is the best approach for the control of infectious diseases. However, conventional screening methods require to operate in well-equipped laboratory. That makes the screening operation expensive and time-consuming. In this work, a reagent-ready-on-chip microfluidic immunoassay system was developed and rapid detection of influenza A H1N1 and H3N2 viruses was demonstrated to show its simplicity and rapidity. The microfluidic system integrated 4 single-stroke thermopneumatic actuators and a reaction chamber. All related reagents were pre-installed in the system and manipulated automatically for indirect immunoassay. Detection of influenza A H1N1 and H3N2 viruses based on its internal structure protein, i.e., nucleoprotein (NP), was demonstrated using the system. Result was represented by colorimetric signal in the reaction chamber. The use of thermopneumatic actuation could achieve fluid manipulation without external assistive equipment and colorimetric result could be observed directly without the need of external dedicated detector. Influenza virus screening could be completed around 1 hour with less sample volume (20 μL) and reagent volume (50 μL). The system achieved one-step operation of immunoassay. It is highly integrated and has potential to be developed to portable devices to realize rapid diagnostics in remote environment and clinics without well-equipped facility. Development of effective disease screening method is the best approach for the control of infectious diseases. However, conventional screening methods require to operate in well-equipped laboratory. That makes the screening operation expensive and time-consuming. In this work, a reagent-ready-on-chip microfluidic immunoassay system was developed and rapid detection of influenza A H1N1 and H3N2 viruses was demonstrated to show its simplicity and rapidity. The microfluidic system integrated 4 single-stroke thermopneumatic actuators and a reaction chamber. All related reagents were pre-installed in the system and manipulated automatically for indirect immunoassay. Detection of influenza A H1N1 and H3N2 viruses based on its internal structure protein, i.e., nucleoprotein (NP), was demonstrated using the system. Result was represented by colorimetric signal in the reaction chamber. The use of thermopneumatic actuation could achieve fluid manipulation without external assistive equipment and colorimetric result could be observed directly without the need of external dedicated detector. Influenza virus screening could be completed around 1 hour with less sample volume (20 μL) and reagent volume (50 μL). The system achieved one-step operation of immunoassay. It is highly integrated and has potential to be developed to portable devices to realize rapid diagnostics in remote environment and clinics without well-equipped facility.

Development of Graphene-based Sensors on Paper Substrate for the Measurement of pH Value of Analyte

Chia-Yi Lee,Kin Fong Lei,Shiao-Wen Tsai,Ngan-Ming Tsang 한국바이오칩학회 2016 BioChip Journal Vol.10 No.3

With excellent properties of graphene material, a number of graphene-based devices have been developed with excellent performance in the past decades. However, difficulty of processing graphene material is a hurdle for its practical realization. In this work, we propose to use vacuum filtration process as the major technique to fabricate graphene-based devices. This technique is simple and effective to practically realize graphene-based sensors on paper substrate, which are equipped with the advantages of low cost, simplicity, flexibility, and disposability. In order to show the functionality of the sensor, determination of pH value of analyte was demonstrated by direct measurement of the resistance across the sensor. Results showed that the sensor has the sensitivity of 30.8 Ω/pH and high linearity (R2=0.9282). Since graphene material can be functionalized by various molecules, it is expected that the graphene-based sensor can be further developed to realize more chemical and biological assays with high sensitivity and specificity for remote environment.

Paper-based Cell Culture Microfluidic System

Fang Fang Tao,Xia Xiao,Kin Fong Lei,I-Chi Lee 한국바이오칩학회 2015 BioChip Journal Vol.9 No.2

In the past decades, glass/PDMS-basedmicrofluidic systems have been rapidly developed to provide homogenous and stable microenvironment for culturing cells. Although these excellent demonstrations involve much simplified operations than traditional cell culture protocol, but they are still not readily accessible to untrained personnel and not appropriate to operate in conventional biological laboratories. In this work, cellulose filter papers were used for the substrates of the cell culture microfluidic system, which provides a convenient tool for cell-based assay. A paper was patterned with culture areas and channels by wax printing technique. Medium or tested substance can be passively perfused to the culture areas. Analyses of cyto-compatibility, cell proliferation, cell morphology, and cell chemosensitivity were performed to confirm the possibility of the paper-based system. Theculture system could provide a platform for a wide range of cell-based assays with applications in drug screening and quantitative cell biology. This work demonstrated a paper-based cell culture microfluidic system and the system is inexpensive, disposable, and compatible to the existing culture facility. In the past decades, glass/PDMS-based microfluidic systems have been rapidly developed to provide homogenous and stable microenvironment for culturing cells. Although these excellent demonstrations involve much simplified operations than traditional cell culture protocol, but they are still not readily accessible to untrained personnel and not appropriate to operate in conventional biological laboratories. In this work, cellulose filter papers were used for the substrates of the cell culture microfluidic system, which provides a convenient tool for cell-based assay. A paper was patterned with culture areas and channels by wax printing technique. Medium or tested substance can be passively perfused to the culture areas. Analyses of cyto-compatibility, cell proliferation, cell morphology,and cell chemosensitivity were performed to confirm the possibility of the paper-based system. The culture system could provide a platform for a wide range of cell-based assays with applications in drug screening and quantitative cell biology. This work demonstrateda paper-based cell culture microfluidic system and the system is inexpensive, disposable, and compatible to the existing culture facility.

The Novel Membrane-Type Micro-system to Assess the Bonus Effect of Physiological and Physical Stimuli on Bone Regeneration

Yen-Ching Yang,Qian-Hui Hong,Kin Fong Lei,Alvin Chao-Yu Chen 한국바이오칩학회 2021 BioChip Journal Vol.15 No.3

The periosteal progenitor cell is suitable for bone tissue regeneration duo to its multipotent differentiation in osteogenesis and chondrogenesis. It was found that both physical and physiological stimuli can induce the differentiation of periosteal progenitor cells. However, the combined-effect of these two stimuli is not clear. The imitation of the nature movement—the cyclic tensile strain stimulation and the multiple growth factors producing cells—adipose-derived stem cells (ADSCs) were used as physical and physiological stimuli to investigate the differentiation of rabbit periosteal cells in this study. For this, a new membrane-type micro-system was invented to provide a simple examination platform for both factors in one single system. The specific rectangular culture chamber not only provided two different types of cells to grow separately but also delivered the single axial tensile strain generated in the micro-system to the cells. It was found that application of either physical or physiological stimuli alone was sufficient to induce the differentiation of periosteal cells. The low tensile strain (4, 5, 6 kPa) led to osteogenesis whereas high tensile strain (7 kPa) induced chondrogenesis. Even though the co-culture of ADSCs only induced osteogenic differentiation of periosteal cells, the co-culture of ADSCs to tensile strain treated periosteal cells further strengthened the osteogenic and chondrogenic differentiation potent in low and high tensile strain, respectively. This study provided the pre-clinical evidence of the stem cell therapy and continuous exercise in cell level bone tissue regeneration.

Osteogenesis and Chondrogenesis of Primary Rabbit Periosteal Cells under Non-uniform 2-Axial Tensile Strain

Chih-Hao Chiu,Yun-Wen Tong,Jen-Fang Yu,Kin Fong Lei,Alvin Chao-Yu Chen 한국바이오칩학회 2020 BioChip Journal Vol.14 No.4

Periosteal cells are the major cell sources of skeletal progenitors for fracture callus. In order to promote bone repair and cartilage formation, besides the application of exogenous growth factors, physical stimulation is an alternative approach to guide cell differentiation. Investigation of appropriate conditions is essential for forming bone and cartilage. In this work, a membrane-type micro-system was developed to provide cell culture environment and cell stretching stimulation during culture course. Circular and oval culture wells were designed to respectively generate uniform and non-uniform 2-axial tensile strain for stretching primary rabbit periosteal cells. Cell orientation and differentiation were studied after cycling stretching for 2 days. The cells aligned to the stretching axis with high tensile strain in the oval culture wells; while the cells expressed random orientation in the circular culture wells. Different responses were significantly shown when the cells were respectively stimulated by uniform and non-uniform 2-axial tensile strains. On the other hand, osteogenic differentiation was shown when the cells were under either uniform or non-uniform 2-axial tensile strain. However, only non-uniform 2-axial tensile strain could induce mature osteoblasts. In addition, the result revealed chondrocytes could be differentiated only under a large and nearly single dimensional tensile strain. In summary, differentiation of the periosteal cells is highly influenced by 2-dimensional distribution of the tensile strain. This work provides some in-sights of the control of axial tensile strain for periosteal cell differentiation.