Cost-Effective Droplet Generator for Portable Bio-Applications.

The convenient division of aqueous samples into droplets is necessary for many biochemical and medical analysis applications. In this article, we propose the design of a cost-effective droplet generator for potential bio-chemical application, featuring two symmetric tubes. The new droplet generator revisits the relationship between capillary components and liquid flow rates. The size of generated droplets by prototype depends only on generator dimensions, without precisely needing to control external flow conditions or driving pressure, even when the relative extreme difference in flow rate for generating nL level droplets is over 57.79%, and the relative standard deviation (RSD) of the volume of droplets is barely about 9.80%. A dropper working as a pressure resource is used to verify the rapidity and robustness of this principle of droplet generation, which shows great potential for a wide range of droplet-based applications.

Construction of Very Low-Cost Loop Polymerase Chain Reaction System Based on Proportional-Integral-Derivative Temperature Control Optimization Algorithm and Its Application in Gene Detection.

Real-time polymerase chain reaction (PCR) technology is essential in nucleic acid detection and point-of-care testing (POCT). However, nowadays, the classical qPCR instrument has the deficiency of its bulky volume, high cost, and inconvenience to use; hence, a low-cost and easy-to-use PCR equipment was thus developed consisting of a hardware subsystem as well as a software subsystem based on an improved proportional-integral-derivative (PID) system. The proposed system not only could hold self-setting reaction cycles of temperature rising and falling automatically but also the temperature during the constant temperature stage was regulated steady based on improved temperature control algorithm, which proved its great effect compared with the reaction temperature derived from an infrared thermal imaging camera. The experimental results in gene detection research also could indicate its applicability and stability of our developed PCR system by using the amplification curve analysis, the melting curve analysis, and agarose gel electrophoresis analysis compared with the commercial PCR instrument, which illustrates the great potential application value of the proposed PCR system.

An integrated digital PCR system with high universality and low cost for nucleic acid detection.

Digital PCR is the most advanced PCR technology. However, due to the high price of the digital PCR analysis instrument, this powerful nucleic acid detection technology is still difficult to be popularized in the general biochemistry laboratory. Moreover, one of the biggest disadvantages of commercial digital PCR systems is the poor versatility of reagents: each instrument can only be used for a few customized kits. Herein, we built a low-cost digital PCR system. The system only relies on low-cost traditional flat-panel PCR equipment to provide temperature conditions for commercial dPCR chips, and the self-made fluorescence detection system is designed and optically optimized to meet a wide range of reagent requirements. More importantly, our system not only has a low cost (<8000 US dollars) but also has a much higher universality for nucleic acid detection reagents than the traditional commercial digital PCR system. In this study, several samples were tested. The genes used in the experiment were plasmids containing UPE-1a fragment, TP53 reference DNA, hepatitis B virus DNA, leukemia sample, SARS-COV-2 DNA, and SARS-COV-2 RNA. Under the condition that DNA can be amplified normally, the function of the dPCR system can be realized with simpler and low-price equipment. Some DNA cannot be detected by using the commercial dPCR system because of the special formula when it is configured as the reaction solution, but these DNA fluorescence signals can be clearly detected by our system, and the concentration can be calculated. Our system is more applicable than the commercial dPCR system to form a new dPCR system that is smaller and more widely applicable than commercially available machinery.

Development of a low-cost multi-channel nucleic acid detection PCR instrument and clinical detection application of COVID-19.

Since the COVID-19 outbreak at the end of December 2019, a variety of novel Coronavirus nucleic acid detection methods have been proposed at home and abroad. Because of the disadvantages of most existing PCR instruments on the market such as long reaction time and high cost, this study developed a more timesaving and cheaper two-channel real-time quantitative PCR instrument. In this instrument, a PCR system combining a thermal cycle system and real-time fluorescence quantitative technology was designed. The software system and data processing, optical system, thermal cycle module, and hardware module of the PCR instrument were studied. The low-cost, portable real-time quantitative PCR system has been validated with consistent results compared to Bio-rad CFX Connect. At the same time, the same samples were used for the contract experiment with the hospital instrument, and the amplification result was better than the existing instrument in the hospital.

Construction of dPCR and qPCR integrated system based on commercially available low-cost hardware.

Fluorescent quantitative PCR (qPCR) and digital PCR (dPCR) are two mainstream nucleic acid quantification technologies. However, commercial dPCR and qPCR instruments have a low integration, a high price, and a large footprint. To solve these shortcomings, we introduce a compound PCR system with both qPCR and dPCR functions. All the hardware used in this compound PCR system is commercially available and low-cost, and free software was used to realize the absolute quantification of nucleic acids. The compound PCR provides two working modes. In the qPCR mode, thermal cycling is realized by controlling the reciprocating motion of the x axis. The heating rate is 1.25 °C s-1 and the cooling rate is 1.75 °C s-1. We performed amplification experiments of the PGEM-3zf (+)1 gene. The performance level was similar to commercial qPCR instruments. In the dPCR mode, the heating rate is 0.5 °C s-1 and the cooling rate is 0.6 °C s-1. We performed the UPE-Q gene amplification and used the sequential actions of the two-dimensional mechanical sliders to scan the reaction products and used the method of regional statistics and back-inference threshold to get test results. The result we got was 1208 copies per µL-1, which was similar to expectations.

Low Cost Three-Dimensional Programmed Mini-Pump Used in PCR.

Programmed mini-pumps play a significant role in various fields, such as chemistry, biology, and medicine, to transport a measured volume of liquid, especially in the current detection of COVID-19 with PCR. In view of the cost of the current automatic pipetting pump being higher, which is difficult to use in a regular lab, this paper designed and assembled a three-dimensional programmed mini-pump with the common parts and components, such as PLC controller, motor, microinjector, etc. With the weighting calibration before and after pipetting operation, the error of the pipette in 10 µL (0.2%), 2 µL (1.8%), and 1 µL (5.6%) can be obtained. Besides, the contrast test between three-dimensional programmed mini-pump and manual pipette was conducted with the ORF1ab and pGEM-3Zf (+) genes in qPCR. The results proved that the custom-made three-dimensional programmed mini-pump has a stronger reproducibility compared with manual pipette (ORF1ab: 24.06 ± 0.33 vs. 23.50 ± 0.58, p = 0.1014; pGEM-3Zf (+): 11.83.06 ± 0.24 vs. 11.50 ± 0.34, p = 0.8779). These results can lay the foundation for the functional, fast, and low-cost programmed mini-pump in PCR or other applications for trace measurements.

Battery-Powered Portable Rotary Real-Time Fluorescent qPCR with Low Energy Consumption, Low Cost, and High Throughput.

The traditional qPCR instrument is bulky, expensive, and inconvenient to carry, so we report a portable rotary real-time fluorescent PCR (polymerase chain reaction) that completes the PCR amplification of DNA in the field, and the reaction can be observed in real-time. Through the analysis of a target gene, namely pGEM-3Zf (+), the gradient amplification and melting curves are compared to commercial devices. The results confirm the stability of our device. This is the first use of a mechanical rotary structure to achieve gradient amplification curves and melting curves comparable to commercial instruments. The average power consumption of our system is about 7.6 W, which is the lowest energy consumption for real-time fluorescence quantification in shunting PCR and enables the use of our device in the field thanks to its self-contained power supply based on a lithium battery. In addition, all of the equipment costs only about 710 dollars, which is far lower than the cost of a commercial PCR instrument because the control system through mechanical displacement replaces the traditional TEC (thermoelectric cooler) temperature control. Moreover, the equipment has a low technical barrier, which can suit the needs of non-professional settings, with strong repeatability.

Low-Cost Battery-Powered and User-Friendly Real-Time Quantitative PCR System for the Detection of Multigene.

Real-time polymerase chain reaction (PCR) is the standard for nucleic acid detection and plays an important role in many fields. A new chip design is proposed in this study to avoid the use of expensive instruments for hydrophobic treatment of the surface, and a new injection method solves the issue of bubbles formed during the temperature cycle. We built a battery-powered real-time PCR device to follow polymerase chain reaction using fluorescence detection and developed an independently designed electromechanical control system and a fluorescence analysis software to control the temperature cycle, the photoelectric detection coupling, and the automatic analysis of the experimental data. The microchips and the temperature cycling system cost USD 100. All the elements of the device are available through open access, and there are no technical barriers. The simple structure and manipulation allows beginners to build instruments and perform PCR tests after only a short tutorial. The device is used for analysis of the amplification curve and the melting curve of multiple target genes to demonstrate that our instrument has the same accuracy and stability as a commercial instrument.