Research Paper
Experimental study on the heating/cooling and temperature uniformity performance of the microchannel temperature control device for nucleic acid PCR amplification reaction of COVID-19

https://doi.org/10.1016/j.applthermaleng.2023.120342Get rights and content

Highlights

  • Novel variable thermal conductivity reaction module is proposed.

  • Multiple parallel serpentine microchannel is used in Polymerase Chain Reaction.

  • Effects of thermal conductivity combination are studied.

  • Heating and cooling rates and temperature uniformity is improved obviously.

Abstract

The nucleic acid detection is an effective way for the prevention and control of COVID-19. PCR amplification is an important process in the nucleic acid detection. At present, PCR amplification has the problem of low heating/cooling rates, and poor temperature uniformity. This paper proposes a microchannel temperature control device for the nucleic acid detection. Five groups of parallel serpentine channels are used to increase the cooling rate of the PCR amplification. A gradual thermal conductivity design is applied to the reaction module to increase the temperature uniformity. The experimental results show that the best temperature uniformity is obtained when the materials of the inner and outer layers of the reaction module are copper and aluminum alloys, respectively. The limit and average heating/cooling rate are 7.2, 6.12, 5.52 and 5.28 °C/s, respectively, when the input power of the thermoelectric cooler is 11.07 W/cm2, the temperature and flow rate of the cooling water are 15℃ and 700 ml/min, and the thermal conductivity of the thermal grease is 6 W/(m·K). Compared with the commercial fan-fin cooling method, the limit and average heating/cooling rates are increased by 38.02%, 80.82%, 86.49% and 208.77%, respectively, with the help of microchannel cooling method.

Graphical abstract

A novel temperature control system including the Vairable thermal conductivity module and the microchannel heating and cooling module was proposed for the nucleic acid testing of COVID-19. The performance of temperature uniformity and heating/cooling rates were improved a lot with the help of the temperature control system.

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Introduction

Polymerase chain reaction (PCR) [1] is a molecular biology technique used to amplify specific DNA fragments. PCR amplification reaction consists three stages of heating-denaturation, cooling-annealing, and heat-preservation-extension [2]. Most of the detection time is spent on the heating and cooling processes. The core problem of the PCR amplification is the precise temperature control of the heating and cooling processes [3]. At present, PCR amplification has the problem of low heating/cooling rates, which results in the long nucleic acid detection time about 2–3 h [4], [5]. Currently, there are two categories of PCR thermocycling mechanism: the temporal method and the spatial method. These two mechanisms are corresponding to two PCR devices: PCR microfluidic chip and PCR amplifier. The PCR amplifier uses the temporal method, and the PCR microfluidic chip uses the spatial method.

The PCR microfluidic chip is a PCR reaction device based on the microfluidic technology. It has the advantages of small size, which is conducive to on-site real-time detection. What’s more, this device has the advantages of small thermal mass, low thermal inertia, fast heat transfer. In addition, the advantages of high integration, fully enclosed, and no manual operation can effectively avoid the contamination of the sample [6], [7]. Refs. [8], [9] developed a fast, battery-powered and portable PCR microfluidic chip. The average heating rate of 32 °C/s and cooling rate of 7.5 °C /s were realized. Although the PCR microfluidic chip has a fast detection speed, the insufficiency of difficult manufacturing, high cost, and a small number of test samples made it hard to meet the large-scale rapid detection requirement of COVID-19. In order to solve this problem, recently, a novel “dry” PCR amplification reaction method based on the spatial mechanism were proposed by Abid et al. [10], [11]. In this method, no water baths was involved, and the capillary tubes with DNA samples were put on the tilting device. There spatial regions with different temperature of 94 °C, 58 °C and 72 °C were designed on the tilting device. The capillary tubes were moved programmed in the three regions and the denaturation, annealing, and extension processes were completed. The advantages of no contamination, no leakage, and free from microbial growth could be obtained by this method.

The PCR amplifier is the most used instrument now days. It has the advantages of large scale, high efficiency and low cost. The reaction module of the PCR amplifier can be made into different specifications according to the detection scale, such as 96 wells, 192 wells, 384 wells and even more. However, the reaction module requires multiple heating and cooling processes. The low heating/cooling rates of only 2–3 °C/s extends the detection time [12]. Increasing the heating/cooling rates can shorten the detection time obviously.

At present, the commercial fan-fin heat dissipation method is the mostly used to improve the heating/cooling rate of PCR amplifier. This method has the advantages of simple structure, low manufacturing cost and high reliability. Chen et al. [13] developed a high precise and fast thermal cycling module. It was heated by a thermoelectric cooler and cooled by a fan. The maximum heating rate of 4 °C/s and cooling rate of 3 °C/s. were obtained. Pogfai et al. [14] developed a fast PCR thermal cycle device with two thermoelectric coolers. The residual heat was dissipated by fins and fans. However, the fan-fin heat dissipation method uses the forced air convection heat transfer mechanism. Due to the low convective heat transfer coefficient, the heating/cooling rate of the PCR amplifier is far from reaching the requirement of rapid nucleic acid detection.

Microchannel heat exchanger has a high heat transfer performance due to the high convective heat transfer coefficient and large heat transfer area, and has been widely used in many fields [15], [16], [17], [18]. In the earlier time, the straight microchannel with smooth surface had been widely used due to the simple structure and easy manufacturing [19]. Later, some microstructures or roughness have been applied to the microchannel heat exchanger to improve the heat transfer performance. Fu et al. [20] designed a microchannel with copper foam fin. The heat transfer coefficient and pressure drop were improved by 80% and 1.2–2 times, respectively. Chang et al. [21] reported a microchannel with nanowires coated micro-pin fins. The heat transfer coefficient was enhanced by 84.6% compared to that of the traditional microchannel. Mandev et al. [22] focused on the influence of surface roughness on the natural and forced convection effects. With the increase of heat exchange performance requirements, more and more novel microchannel structures have been proposed. Zheng et al. [15] proposed a zigzag-type microchannel heat exchanger with asymmetric geometry to improve the thermal–hydraulic performance. Al-Neama et al. [23] proposed a serpentine microchannel and applied it to the cooling of microelectronic chips. Because of the excellent heat transfer performance, the microchannel has good application potential in PCR amplification reactions.

Besides the heating/cooling rates, the temperature uniformity is another problem that needs to be solved. Good temperature uniformity improves the quality of amplified DNA. Therefore, the temperature uniformity of PCR amplification reaction has aroused great interest of scholars. Hsieh et al. [24] proposed a new type of array micro heater with active compensation unit, and the temperature uniformity was obviously improved. When the reaction temperatures were 94 °C, 57 °C and 72 °C, respectively, the uniform temperature area that the temperature variation less than 1 °C reached 63.6%, 96.6% and 79.6%. Imran et al. [25] introduced 0.5 mm, 1 mm, and 1.5 mm air gaps in the glass substrate to improve the temperature uniformity in the PCR reaction. Chen et al. [26] developed a PCR chip with PMMA microchannel and aluminum cover. When the air gap and thickness of the microchannel were both 1 mm, the temperature uniformity was increased obviously.

According to the above reports, the most used temperature control scheme of the PCR amplifier is the combination of thermoelectric cooler, fins, and fan. This method has the disadvantage of low heating/cooling rates. The temperature uniformity is mainly controlled by the method of temperature compensation. This method has the disadvantage of high manufacturing and maintenance costs, and the service life is difficult to guarantee.

In order to improve the heating/cooling rates and temperature uniformity, in this paper, a microchannel temperature control method for PCR amplification reaction is proposed. The microchannel contains five serpentine channels. Two flow distribution channels are located at the inlet and outlet of the microchannel. The cooling rate and temperature uniformity are improved simultaneously. In addition, the reaction module is designed with a gradual thermal conductivity, and this can further improve the temperature uniformity. The influence of input power of thermoelectric cooler, the temperature and flow rate of the cooling water, and the thermal conductivity combination of the reaction module on the heating and cooling rates of the PCR amplification reaction as well as the temperature uniformity are studied.

Section snippets

Temperature control system for PCR amplification reaction

At present, the most used DNA amplification equipment in nucleic acid detection is the PCR amplifier. It combines the functions of nucleic acid amplification, large-scale detection and analysis, and has made an important contribution to the prevention and control of COVID-19. The most important process of the PCR amplification reaction is the temperature control. The temperature control system of the PCR amplification reaction is shown in Fig. 1 (a), including the reaction module, the heating

Temperature uniformity

Temperature uniformity is an important performance for the PCR amplification reaction. In order to achieve the rapid nucleic acid detection, multiple nucleic acid samples are often amplified in a reaction module. The number of samples in a reaction module is generally 24, 48, 96, 192, 384 or even more. During the heating and cooling processes, in order to make sure that the amplified samples have the same DNA concentration, it is necessary to keep the same temperature for each sample.

Conclusions

In this research, a PCR reaction module with gradual thermal conductivity and multiple parallel serpentine microchannel temperature control method are proposed. The temperature uniformity and heating and cooling rates of the PCR amplification reaction are effectively improved by optimizing the parameters of the reaction module. The conclusions are as follows:

  • (1)

    The PCR reaction module are divided into an inner layer and an outer layer. Four materials including copper, aluminum alloy, brass and

Declaration of Competing Interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Acknowledgement

This work was supported by the National Natural Science Foundation of China (No. 52205497), the Natural Science Foundation of Xiamen (No. 3502Z20227027), the Fundamental Research Funds for the Central Universities (No. 20720220092), and the Major Science and Technology Program of Xiamen City (No. 3502Z20231009). In addition, the support from the Higher Educational Key Laboratory for Flexible Manufacturing Equipment Integration of Fujian province (Xiamen Institute of Technolog) and the

References (30)

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