Rtca dp analyzer

HK-2 cells (4 × 10⁵ cells/6-cm culture dish) were grown overnight and treated at the specified conditions for 72 h. Cells were then collected, washed with PBS (3 mL), and centrifuged at 200 × g for 5 min. The xCELLigence system (RTCA DP Analyzer, Roche Applied Science, Penzberg, Germany) was used for cell analysis. The system consisted of RTCA Resistor Plate 16 devices (ACEA Biosciences Inc., San Diego, CA, USA) placed in an incubator system (humidified atmosphere 5% CO₂ at 37 °C; Thermo Fisher Scientific Inc.) and RTCA DP software (version 1.2) that was used to verify system conditions and for sample monitoring. Wells of the E-Plate 16 devices (ACEA Biosciences Inc.) were filled with 100 μL of RPMI 1640 media (containing 10% FBS and 1% penicillin/streptomycin), and the background value was measured using the RTCA DP software. Subsequently, harvested cells were counted using a hemocytometer (Paul Marienfeld GmbH & Co.), and cells (1 × 105 (link) cells/well in 100 μL RPMI 1640 media) were seeded on E-Plate 16 devices connected to the xCELLigence system. Real-time cell swelling and adhesion were measured using the RTCA DP software at 10 s intervals and monitored for 3 h. The real-time cell index was determined using the RTCA DP software, and a histogram representative of the cell index for 3 h was generated.

MDA-MB-231 cells were seeded at 20,000 cells per well in serum-free 1640 medium with or without 50 ng/mL IL-6 in the upper chamber of CIM-plate16. And 150 µL 1640 medium with 10% FBS was added in the lower chamber of CIM-plate16. Cells were then treated with PN. Insert the CIM-plate 16 into the RTCA DP Analyzer (Roche Applied Science, Penzberg, Germany). The following steps were performed as described [50 (link)].

For monitoring the effects of nanoparticles on HUVEC viability, a xCELLigence system (RTCA DP Analyzer, Roche Diagnostics, Mannheim, Germany) was used. Experiments were performed in 16-well E-plates containing microelectrodes for impedance measurement (ACEA Bioscience, San Diego, CA, USA).
For the background measurement, 100 µL of cell-free endothelial cell growth medium was added to the wells. Afterwards, 50 µL of media from each well were replaced with 50 µL of cell suspension containing 1 × 10³ HUVECs and monitoring of impedance was initiated. At 24 h after seeding, an additional 100 µL of media containing WPI at concentrations 2x higher than the required final concentrations was added. The final WPI concentrations were as follows: 0, 50, 150 and 500 µg/mL. Cell growth was monitored every 10 min for 96 h. The experiments were performed in hexaplicate.

Real-time cell migration monitoring was conducted using the xCELLigence system (Roche Applied Science, Penzberg, Upper Bavaria, Germany) following the manufacturer’s protocol. In brief, 8x10⁴ cells were seeded into each well in the upper chamber of the CIM plate, which was coated with collagen. The lower chamber containing 10% serum media attracts the cells across the chambers, which will then be recorded real-time using a RTCA DP analyzer (Roche Applied Science, Penzberg, Upper Bavaria, Germany).

NMuMG-ErbB2 cells (5 × 10³) and 1 × 10⁴ NIC cells were plated into xCELLigence E-plates (Cat. #: 05469813001, Roche Applied Science) and incubated in the presence or absence of TGFβ. Cell growth was monitored in a RTCA DP Analyzer (Roche Applied Science) for up to 96 h and the doubling time was calculated using the xCELLigence RTCA software (Roche Applied Science) according to manufacturer's protocol.