The culture of A549 cells and its secreted cytokine IL-6 monitoring on the designed multifunctional microfluidic chip.

Abstract

A multifunctional microfluidic chip integrated with perfusion cell culture and in situ SERS detection of cell secretion was designed and developed for the detection of IL-6 secretion from LPS-stimulation of A549 cells in this paper. Researching works were focused on A549 cell activity and secretion in the constructed LPS-stimulated A549 cells model. On the designed microchip, a bubble trap chamber was designed to remove the bubbles in the culture medium which could also be simultaneously preheated by a split hot plate. Then, a long-time perfusion culture process of 549 cells could be realized. Under the optimized conditions the A549 cells could be cultured and kept in good activity for more than 36 h. Subsequently, the model of interaction between LPS and A549 cells was established on the designed microchip. When LPS-stimulated A549 cells, the IL-6 which was one of the secretions formed in this process was detected quantitatively by SERS spectral technique. The silver-coated gold nano-stars were prepared and taken as a sensitive enhancing probe for the SERS detection of IL-6 secreted from LPS-stimulated A549 cells. The immunomagnetic beads, IL-6 antigen, and SERS probes were mixed and incubated in the microchip and form a sandwich structure which was captured by the permanent magnet in the detection zone for SERS detection. The reference material of IL-6 was used to establish the calibration curve, and the linear range and detection limit were 1-10000 pg/mL and 0.75 pg/mL, respectively. Then, the IL-6 secretion from LPS-stimulated A549 cells was detected hourly for 7 h by this established method. The process of LPS stimulation of A594 cells did not lead to a sustained increase in the SERS spectral signature of IL-6. Instead, IL6 secretion initially increased sharply, then decreased and eventually stabilized. It could be due to a potential mechanism that the cells self-regulated to mitigate the inflammatory effects in response to sustained stimulation. The proposed multifunctional microfluidic chip, characterized by high sensitivity and the ability to perform continuous hourly detection, exhibited significant application prospects in the study of external stimulation on cells.