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Application of optically-induced-dielectrophoresis in microfluidic system for purification of circulating tumour cells for gene expression analysis- Cancer cell line model

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ABSTRACT

Circulating tumour cells (CTCs) in a blood circulation system are associated with cancer metastasis. The analysis of the drug-resistance gene expression of cancer patients’ CTCs holds promise for selecting a more effective therapeutic regimen for an individual patient. However, the current CTC isolation schemes might not be able to harvest CTCs with sufficiently high purity for such applications. To address this issue, this study proposed to integrate the techniques of optically induced dielectrophoretic (ODEP) force-based cell manipulation and fluorescent microscopic imaging in a microfluidic system to further purify CTCs after the conventional CTC isolation methods. In this study, the microfluidic system was developed, and its optimal operating conditions and performance for CTC isolation were evaluated. The results revealed that the presented system was able to isolate CTCs with cell purity as high as 100%, beyond what is possible using the previously existing techniques. In the analysis of CTC gene expression, therefore, this method could exclude the interference of leukocytes in a cell sample and accordingly contribute to higher analytical sensitivity, as demonstrated in this study. Overall, this study has presented an ODEP-based microfluidic system capable of simply and effectively isolating a specific cell species from a cell mixture.

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Evaluation of the maximum manipulation velocities and ODEP manipulation forces generated under different voltage and light image conditions: (a) voltage magnitude range: 2–10 V, bandwidth of long rectangular light bar: 150, 200, and 250 μm; (b) voltage magnitude 2–10 V, bandwidth of hollow circular light images (fixed ID: 40 μm): 20, 40, and 60 μm [results given as the mean ± standard deviation of 3 separate experiments (n = 9)]. (ODEP+: positive ODEP; ODEP−: negative ODEP force).
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f4: Evaluation of the maximum manipulation velocities and ODEP manipulation forces generated under different voltage and light image conditions: (a) voltage magnitude range: 2–10 V, bandwidth of long rectangular light bar: 150, 200, and 250 μm; (b) voltage magnitude 2–10 V, bandwidth of hollow circular light images (fixed ID: 40 μm): 20, 40, and 60 μm [results given as the mean ± standard deviation of 3 separate experiments (n = 9)]. (ODEP+: positive ODEP; ODEP−: negative ODEP force).

Mentions: Figure 4(a) revealed the effect of the voltage magnitude applied (2–10 V) and the bandwidth of the light bar utilized (150, 200, and 250 μm) on the measured maximum velocities of the light bar (and the corresponding manipulation force calculated: right y axis) that can manipulate the live and dead leukocytes. For both a positive and negative ODEP (ODEP+ and ODEP−) force, the results in Fig. 4(a) corresponded well with equation (2), showing that the magnitude of the maximum manipulating velocity (or manipulation force) was proportional to the voltage squared. Although a higher magnitude of electric voltage could correspondingly generate a higher ODEP manipulation force for more efficient cell manipulation, ODEP force-based cell manipulation under a high electric field could lead to cell aggregation, which in turn might affect the cell separation process described in Fig. 2. This phenomenon was also observed and discussed in previous studies36. In this study, the magnitude of 8 V was used, which was a compromise between the manipulation force generated for cell manipulation and the cell aggregation phenomenon based on our microscopic observations. Regarding the influence of the bandwidth of the light bar on the ODEP manipulation forces, at a given voltage, the condition of 150 μm bandwidth was measured to have a statistically higher magnitude of maximum manipulating velocity (or manipulation force) than 250 μm bandwidth in the cases of both live and dead leukocytes [Fig. 4(a)]. This finding could be explained by the fact that, under an ODEP field, the electric field within a smaller light image could be more focused than in a larger one, accordingly contributing to a higher ODEP manipulation force44. Therefore, the light bar with 150 μm bandwidth was used for leukocyte manipulation in this work. After the magnitude of the electric voltage (8 V) and bandwidth of the light bar (150 μm) were determined, the maximum velocities of the moving light bar that can manipulate (pull) live and (push) dead leukocytes were measured to be 250.49 ± 34.63 μm/s and 115.57 ± 7.73 μm/s, respectively [Fig. 4(a)]. In the operation processes described in Fig. 2, a long moving rectangular light bar was utilized to sweep all the leukocytes to one side of the main microchannel for separation from the target cancer cells. For this purpose, the moving velocity of the light bar was set to 100 μm/s, allowing all the live and dead leukocytes to be simultaneously pulled and pushed, respectively, to one side of the main microchannel based on the above experimental evaluations.


Application of optically-induced-dielectrophoresis in microfluidic system for purification of circulating tumour cells for gene expression analysis- Cancer cell line model
Evaluation of the maximum manipulation velocities and ODEP manipulation forces generated under different voltage and light image conditions: (a) voltage magnitude range: 2–10 V, bandwidth of long rectangular light bar: 150, 200, and 250 μm; (b) voltage magnitude 2–10 V, bandwidth of hollow circular light images (fixed ID: 40 μm): 20, 40, and 60 μm [results given as the mean ± standard deviation of 3 separate experiments (n = 9)]. (ODEP+: positive ODEP; ODEP−: negative ODEP force).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5016898&req=5

f4: Evaluation of the maximum manipulation velocities and ODEP manipulation forces generated under different voltage and light image conditions: (a) voltage magnitude range: 2–10 V, bandwidth of long rectangular light bar: 150, 200, and 250 μm; (b) voltage magnitude 2–10 V, bandwidth of hollow circular light images (fixed ID: 40 μm): 20, 40, and 60 μm [results given as the mean ± standard deviation of 3 separate experiments (n = 9)]. (ODEP+: positive ODEP; ODEP−: negative ODEP force).
Mentions: Figure 4(a) revealed the effect of the voltage magnitude applied (2–10 V) and the bandwidth of the light bar utilized (150, 200, and 250 μm) on the measured maximum velocities of the light bar (and the corresponding manipulation force calculated: right y axis) that can manipulate the live and dead leukocytes. For both a positive and negative ODEP (ODEP+ and ODEP−) force, the results in Fig. 4(a) corresponded well with equation (2), showing that the magnitude of the maximum manipulating velocity (or manipulation force) was proportional to the voltage squared. Although a higher magnitude of electric voltage could correspondingly generate a higher ODEP manipulation force for more efficient cell manipulation, ODEP force-based cell manipulation under a high electric field could lead to cell aggregation, which in turn might affect the cell separation process described in Fig. 2. This phenomenon was also observed and discussed in previous studies36. In this study, the magnitude of 8 V was used, which was a compromise between the manipulation force generated for cell manipulation and the cell aggregation phenomenon based on our microscopic observations. Regarding the influence of the bandwidth of the light bar on the ODEP manipulation forces, at a given voltage, the condition of 150 μm bandwidth was measured to have a statistically higher magnitude of maximum manipulating velocity (or manipulation force) than 250 μm bandwidth in the cases of both live and dead leukocytes [Fig. 4(a)]. This finding could be explained by the fact that, under an ODEP field, the electric field within a smaller light image could be more focused than in a larger one, accordingly contributing to a higher ODEP manipulation force44. Therefore, the light bar with 150 μm bandwidth was used for leukocyte manipulation in this work. After the magnitude of the electric voltage (8 V) and bandwidth of the light bar (150 μm) were determined, the maximum velocities of the moving light bar that can manipulate (pull) live and (push) dead leukocytes were measured to be 250.49 ± 34.63 μm/s and 115.57 ± 7.73 μm/s, respectively [Fig. 4(a)]. In the operation processes described in Fig. 2, a long moving rectangular light bar was utilized to sweep all the leukocytes to one side of the main microchannel for separation from the target cancer cells. For this purpose, the moving velocity of the light bar was set to 100 μm/s, allowing all the live and dead leukocytes to be simultaneously pulled and pushed, respectively, to one side of the main microchannel based on the above experimental evaluations.

View Article: PubMed Central - PubMed

ABSTRACT

Circulating tumour cells (CTCs) in a blood circulation system are associated with cancer metastasis. The analysis of the drug-resistance gene expression of cancer patients’ CTCs holds promise for selecting a more effective therapeutic regimen for an individual patient. However, the current CTC isolation schemes might not be able to harvest CTCs with sufficiently high purity for such applications. To address this issue, this study proposed to integrate the techniques of optically induced dielectrophoretic (ODEP) force-based cell manipulation and fluorescent microscopic imaging in a microfluidic system to further purify CTCs after the conventional CTC isolation methods. In this study, the microfluidic system was developed, and its optimal operating conditions and performance for CTC isolation were evaluated. The results revealed that the presented system was able to isolate CTCs with cell purity as high as 100%, beyond what is possible using the previously existing techniques. In the analysis of CTC gene expression, therefore, this method could exclude the interference of leukocytes in a cell sample and accordingly contribute to higher analytical sensitivity, as demonstrated in this study. Overall, this study has presented an ODEP-based microfluidic system capable of simply and effectively isolating a specific cell species from a cell mixture.

No MeSH data available.


Related in: MedlinePlus