Non-Interventional and High-Precision Temperature Measurement Biochips for Long-Term Monitoring the Temperature Fluctuations of Individual Cells

Abstract

Monitoring the thermal responses of individual cells to external stimuli is essential for studies of cell metabolism, organelle function, and drug screening. Fluorescent temperature probes are usually employed to measure the temperatures of individual cells; however, they have some unavoidable problems, such as, poor stability caused by their sensitivity to the chemical composition of the solution and the limitation in their measurement time due to the short fluorescence lifetime. Here, we demonstrate a stable, non-interventional, and high-precision temperature-measurement chip that can monitor the temperature fluctuations of individual cells subject to external stimuli and over a normal cell life cycle as long as several days. To improve the temperature resolution, we designed temperature sensors made of Pd-Cr thin-film thermocouples, a freestanding Si₃N₄ platform, and a dual-temperature control system. Our experimental results confirm the feasibility of using this cellular temperature-measurement chip to detect local temperature fluctuations of individual cells that are 0.3-1.5 K higher than the ambient temperature for HeLa cells in different proliferation cycles. In the future, we plan to integrate this chip with other single-cell technologies and apply it to research related to cellular heat-stress response.

Keywords: biochip fabrication; cell metabolism; cell temperature; non-interventional biosensors; single-cell temperature measurement; temperature sensors.

Figure 1

Schematic illustration of the new biochip and measurement system. (a) Schematic diagram of a cross-section of the functional area on the chip. The cream-colored bars labeled PDMS represent a cross-section of the polydimethylsiloxane ring that contains the cell-culture pool (see Experimental Section). (b) Optical photo of the whole chip, including the functional area. The chip is mounted on a printed-circuit board (PCB). (c) The working principle of the homemade, low-noise cell-temperature detection system.

Figure 2

Effects of surface modifications of the chips. (a) Photos of the 48- and 72-h morphologies of HeLa cells on chips with different surface treatments. (b) Cell-temperature data collected from the unprocessed chip and from chips with poly-l-lysine and polydopamine modifications.

Figure 3

The high thermal sensitivity and low thermal noise of the cellular temperature-monitoring chip and system. (a) Background-temperature curves from the developed chips. (b) Comparing the output thermoelectric signals from the temperature sensors on freestanding Si₃N₄ platform and on solid Si₃N₄/Si/Si₃N₄ substrate exposed to a HeLa cell culture and to a blank control group without cells shows the high thermal sensitivity of the chips. (c) Enlarged temperature curves of HeLa cells obtained from the sensors on the Si₃N₄/Si/Si₃N₄ substrates show a low temperature sensitivity. (d) Thermal-noise test results for three standard commercial thermocouples. (e) Thermal-noise test results for the developed chips with Pd–Cr TFTC sensors on a freestanding Si₃N₄ platform.

Figure 4

(a–d) The absolute temperature fluctuations of the HeLa cells on the four TFTC sensors named 1Au2, 1Au7, 1Cu8, and 1Cd2 in a 37 °C environment after removal of the background thermal noise. The gray curves represent the original measurement data, and the orange, purple, green, and dark red thick curves result from smoothing the original data. (e,f) Photos of the relative positions of the under-test cells and the four thermocouple sensors are highlighted by red dots.

Figure 5

The temperatures of the HeLa cells drop sharply upon the addition of a lethal fluid (the red arrows represents the moment when concentrated HCl is injected into the medium). (a) Two typical curves showing the temperature changes caused by the deaths of the HeLa cells due to the addition of concentrated HCl. (b) The control group shows no obvious temperature drop in the HeLa cells on the Si₃N₄/Si/Si₃N₄ substrate when subjected to concentrated HCl. (c) The blank control group containing the culture medium without HeLa cells also shows no obvious temperature drop after the injection of concentrated HCl. (d) Temperature changes of the culture medium caused by the exothermic effects of concentrated H₂SO₄ and of concentrated HCl. (e) The morphology of HeLa cells at the moments 1 min, 2 min, and 5 min after the introduction of concentrated HCl.