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Results: 7

1.
Fig. 5

Fig. 5. DEP exposure does not interfere with differentiation of NSPCs into neuronal and astrocytic lineages. From: Advancing practical usage of microtechnology: a study of the functional consequences of dielectrophoresis on neural stem cells.

(A) Schematic portrays the experimental design to test effects of DEP on neuronal and astrocytic lineage potential of NSPCs. (B) Left panel: The neuronal lineage potential of SC27 HuNSPCs (as shown by generation of MAP2+ cells) is not altered by long-term DEP exposure at various DEP frequencies. Control cells were not exposed to DEP force. Right panel: Image shows HuNSPCs (SC27) differentiated for 14 days and stained with MAP2 antibody (red) to detect neurons. All nuclei are stained blue. (C) Left panel: The astrocytic lineage potential of SC27 HuNSPCs (as shown by generation of GFAP+/Sox2− cells) is not modified by long-term DEP exposure at various DEP frequencies. Right panel: Image shows HuNSPCs (SC27) differentiated for 7 days and co-stained with GFAP (red) and Sox2 (green) antibodies (GFAP+/Sox2− astrocytes are shown by arrows). All nuclei are stained blue. Data are represented as mean ± SE.

Jente Lu, et al. Integr Biol (Camb). 2012 October;4(10):10.1039/c2ib20171b.
2.
Fig. 4

Fig. 4. DEP exposure does not alter NSPC proliferation. From: Advancing practical usage of microtechnology: a study of the functional consequences of dielectrophoresis on neural stem cells.

(A) Schematic depicts the experimental design to test the effects of DEP on cell proliferation. (B) Left panel: Image of untreated HuNSPCs (SC27) demonstrates that 5–10 % of the cells are EdU-positive (red). All nuclei are stained blue. Right panel: No significant difference in cell division as measured by EdU incorporation was observed after DEP exposure for 1 or 30 min at all frequencies tested. (C) Control cells and those exposed to various DEP frequencies did not significantly differ in cell cycle kinetics. Cells were analyzed immediately after DEP exposure and at various times to allow up to 2 cell cycles to occur (~48 h doubling time). Data are represented as mean ± SE.

Jente Lu, et al. Integr Biol (Camb). 2012 October;4(10):10.1039/c2ib20171b.
3.
Fig. 3

Fig. 3. Long-term DEP exposure to frequencies of 50 kHz and 100 kHz, but not lower or higher frequencies, decreases cell survival. From: Advancing practical usage of microtechnology: a study of the functional consequences of dielectrophoresis on neural stem cells.

(A) Survival of HuNSPCs (SC27) as determined by trypan blue exclusion after DEP exposure is most affected by 50 and 100 kHz frequencies. (B) The cell survival of three different sets of NSPCs (SC23 or SC27 HuNSPCs and E12.5 mNSPCs) is decreased after 30 min exposure to DEP frequencies of 50 kHz and 100 kHz, but not other frequencies. (C) Left panel: Amount of LDH released by control cells (in DEP buffer but not exposed to DEP force) is set as 0% LDH release and amount of LDH released by cells that were completely lysed with triton-X100 is set as 100% LDH release. Right panel: Release of LDH by cells after exposure to DEP frequencies is greatest at 50 kHz. (D) Cellular metabolic activity was decreased primarily by long-term DEP exposure to frequencies of 50 and 100 kHz. Data are represented as mean ± SE (*, p-value < 0.05 and **, p-value <0.01).

Jente Lu, et al. Integr Biol (Camb). 2012 October;4(10):10.1039/c2ib20171b.
4.
Fig. 1

Fig. 1. Multi-well device and experimental setup for testing effects of DEP force on NSPCs. From: Advancing practical usage of microtechnology: a study of the functional consequences of dielectrophoresis on neural stem cells.

(A) DEP exposure well (top schematic) contains interdigitated gold-plated electrodes at the bottom of the well (sides of well are PDMS). Fifteen individual wells (red square) make up the multi-well DEP device (bottom schematic), which is connected to a function generator to energize the electrodes (energized by 8Vpeak-peak). The bottom electrical head serves as a DEP force switch for each group of three exposure wells (blue square). The symbol, , represents a switch in the circuit. (B) Undifferentiated HuNSPCs (SC27) are grown as adherent cultures (phase contrast panel) and express the neural stem cell markers Sox2 and Nestin (fluorescence panel, 98.0 +/− 0.5% of the cells express either Sox2 or Nestin and 78.6 +/− 2.7% co-express both markers, N=1300 cells). All cell nuclei are stained with Hoechst. NSPCs are grown as adherent cultures but were dissociated to form a cell suspension for all DEP exposure experiments. Data are represented as mean ± SE.

Jente Lu, et al. Integr Biol (Camb). 2012 October;4(10):10.1039/c2ib20171b.
5.
Fig. 2

Fig. 2. Short-term DEP exposure has negligible effects on NSPC survival. From: Advancing practical usage of microtechnology: a study of the functional consequences of dielectrophoresis on neural stem cells.

(A) Schematic of experimental design to test effects of short-term and long-term DEP exposure on cell survival. (B) HuNSPCs exposed to DEP frequencies for 0, 10, or 60 sec exhibit no significant decrease in cell survival as determined by trypan blue exclusion. (C) The cell survival of three different sets of NSPCs (HuNSPCs SC23 or SC27 and mNSPCs E12.5) is not affected by 60 sec DEP exposure at any frequency. (D) Left panel: The baseline level of LDH released by intact control HuNSPCs (SC27 cells in DEP buffer not exposed to DEP force) is set as 0%. The maximum amount of LDH release is shown for HuNSPCs (SC27) lysed with triton-X100 and is set as 100% LDH released. Right panel: Release of LDH by cells after 60 sec exposure to DEP frequencies is only above background levels at 50 kHz. (E) The cellular metabolic activity of HuNSPCs (SC27) after 60 sec exposure to DEP frequencies remains greater than 90% for all conditions. Data are represented as mean ± SE (*, p-value < 0.05 and **, p-value <0.01).

Jente Lu, et al. Integr Biol (Camb). 2012 October;4(10):10.1039/c2ib20171b.
6.
Fig. 7

Fig. 7. Electrode and gap size optimization for maximal cell deflection and minimal electric field exposure. From: Advancing practical usage of microtechnology: a study of the functional consequences of dielectrophoresis on neural stem cells.

(A) The left panel schematic depicts an example of the CFD modeling used to determine the deflection of a particle in response to the DEP electric field from a pair of interdigitated electrodes set to 8V peak-peak. Particles and fluid enter the channel from the left and as the particle passes by the electrode and gap it is deflected upon encountering the electric field. Delta Y represents the distance the particle moved due to the induced DEP force. The right panel color surface plot shows delta Y as a function of the sizes of the electrodes and the gaps between them. (B) The left panel shows a color plot of the electric field distribution above the electrodes for a 50 μm electrode × 100 μm gap configuration. One pair of electrodes with intervening gap is enlarged in the dashed box to demonstrate the area used to calculate the mean electric field for each electrode/gap (blue and red box), which is graphed in the right panel. The right panel graph portrays the mean electric field, which is proportional to the induced transmembrane potential (ΔU), for multiple electrode and gap sizes and combinations.

Jente Lu, et al. Integr Biol (Camb). 2012 October;4(10):10.1039/c2ib20171b.
7.
Fig. 6

Fig. 6. Rapid increases in the transmembrane potential of cells at frequencies slightly above the crossover frequency cause membrane disruption of NSPCs in suspension. From: Advancing practical usage of microtechnology: a study of the functional consequences of dielectrophoresis on neural stem cells.

(A) The iso-curves represent the root mean square of the gradient of the electric field square in the z direction (see equation above graph). The induced DEP force is proportional to the iso-curve value. The red and green boxes represent electrodes with either positive or negative applied voltages. (B) Graph shows the simulated CM factor (parameters in Table S1 in Supporting Material) of HuNSPCs (SC27) at various frequencies. The blue diamonds represent the CM factor of SC27 cells at frequencies used in the current study to test the toxicity of DEP exposure to cells. The derived crossover frequency of SC27 cells is 45 kHz (red dashed line). (C) Schematic represents cells’ final positions relative to electrodes at various applied frequencies. (D) Time lapse images show behavior of a cell (circled in green) in response to 50 kHz DEP frequency over ~2.4 sec. The dashed black line represents the edge of the electrode (high electric field) while the solid blue line denotes the position of cells that have moved away from the electrode (lower electric field). (E) The graph shows the induced transmembrane potential of a hypothetical population of adherent SC27 cells at the edge of electrodes. The lower the frequency applied, the higher the induced transmembrane potential of NSPCs. The red dashed line represents the resting membrane potential of SC27 HuNSPCs, 51.6+/−2.6 mV, n=20. The green dashed line represents the crossover frequency of SC27 cells (~45 kHz). The parameters for the modeling are listed in Table S1 in Supporting Material. (F) The graph depicts the frequency-dependent induced transmembrane potential of suspended SC27 cells accounting for the final position of the cells relative to the electrode (and thus the strength of the electric field). The black line denotes the induced transmembrane potential for cells in a region of high electric field near the electrodes, as shown by the black dashed line in D, and the blue line denotes the induced transmembrane potential for cells in a region of low electric field farther from the electrodes, as shown by the blue line in D. The parameters for the modeling are listed in Table S1 in Supporting Material.

Jente Lu, et al. Integr Biol (Camb). 2012 October;4(10):10.1039/c2ib20171b.

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