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Wang, Z.; Sun, M.; Zhang, Q.; Li, P.; Wang, K.; Li, X. Point-of-Care Testing of microRNAs. Encyclopedia. Available online: https://encyclopedia.pub/entry/47573 (accessed on 07 October 2024).
Wang Z, Sun M, Zhang Q, Li P, Wang K, Li X. Point-of-Care Testing of microRNAs. Encyclopedia. Available at: https://encyclopedia.pub/entry/47573. Accessed October 07, 2024.
Wang, Zhong-Yu, Ming-Hui Sun, Qun Zhang, Pei-Feng Li, Kun Wang, Xin-Min Li. "Point-of-Care Testing of microRNAs" Encyclopedia, https://encyclopedia.pub/entry/47573 (accessed October 07, 2024).
Wang, Z., Sun, M., Zhang, Q., Li, P., Wang, K., & Li, X. (2023, August 02). Point-of-Care Testing of microRNAs. In Encyclopedia. https://encyclopedia.pub/entry/47573
Wang, Zhong-Yu, et al. "Point-of-Care Testing of microRNAs." Encyclopedia. Web. 02 August, 2023.
Point-of-Care Testing of microRNAs
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MicroRNAs (miRNAs) are a class of small noncoding RNAs that are approximately 22 nt in length and regulate gene expression post-transcriptionally. miRNAs play a vital role in both physiological and pathological processes and are regarded as promising biomarkers for cancer, cardiovascular diseases, neurodegenerative diseases, and so on. Accurate detection of miRNA expression level in clinical samples is important for miRNA-guided diagnostics. However, the common miRNA detection approaches like RNA sequencing, qRT-PCR, and miRNA microarray are performed in a professional laboratory with complex intermediate steps and are time-consuming and costly, challenging the miRNA-guided diagnostics. Hence, sensitive, highly specific, rapid, and easy-to-use detection of miRNAs is crucial for clinical diagnosis based on miRNAs. With the advantages of being specific, sensitive, efficient, cost-saving, and easy to operate, point-of-care testing (POCT) has been widely used in the detection of miRNAs.

microRNA (miRNA) point-of-care testing (POCT) visual detection portable instruments

1. Introduction

MicroRNAs (miRNAs) are a type of small noncoding RNA with a length of ~21–25 nt that act as regulators of gene expression at the post-transcriptional level [1]. The miRNA genes are transcribed into hairpin-containing pre-miRNA by RNA polymerase III, and the long dsRNA precursors are processed by Drosha and Dicer consecutively [2][3]. The generated small dsRNAs are loaded onto an argonaute family protein (AGO) to form an RNA-induced silencing complex (RISC). After loading, the passenger strand of the miRNA duplex exits to produce a single-stranded mature miRNA, and the mature RISC induces translational repression, mRNA deadenylation, and mRNA decay [4][5]. miRNAs play vital roles in development. miRNAs regulate cellular activities, including cell growth, differentiation, and apoptosis, and aberrant expression of miRNAs promotes the occurrence and development of diseases. In recent decades, miRNAs have been implicated in various human diseases. Hence, many studies have attempted to apply miRNAs to disease diagnosis, and miRNAs show great promise as diagnostic biomarkers, as miRNAs can not only circulate in the human blood in remarkably stable forms, such as exosomes, but they are also widely present in other bio-microenvironments, such as urine, saliva, and cerebrospinal fluid [6][7]. Accurate detection of dysregulated circulating miRNAs in biofluids is important for miRNA-guided diagnostics in a noninvasive fashion. There have been many conventional methods for the quantitative detection of miRNAs, such as northern blot, microarray, RNA-seq and RT-qPCR [8]. Although these traditional methods are relatively highly sensitive and specific, these approaches also have various limitations. For example, northern blotting and real-time PCR are sensitive and specific, but they are also labor-intensive and require specialized equipment. Microarray and RNA-seq are high-throughput methods that allow the simultaneous detection of multiple miRNAs, but they are also expensive and require complex data analysis, and these approaches for miRNA detection are performed in a professional laboratory, which is challenging for the application of miRNA detection in clinical practice. Therefore, it has driven the development of reliable point-of-care testing (POCT) of miRNAs. Point-of-care testing (POCT) is defined as testing performed near or in the field of a patient, for whom faster results may lead to changes in patient care [9]. Recently, POCT has been applied to the quantitative detection of miRNAs and has made rapid progress. To be more detailed, POCT can provide accurate and ultrasensitive tumor screening results for patients with the advantages of a no-fuss operation, low cost, and rapidity [10][11][12][13]. At the same time, POCT is also suitable for resource-limited areas, or even for self-testing. Previous reviews provided valuable information on the evolution of POCT-detection methods for miRNAs and the applied amplification strategies in POCT for miRNAs [14][15]. The development in detection of multiple miRNAs and the new progress in biosensors, microfluidics, and lateral flow assays (LFAs) for miRNA detection have also been well reviewed [16][17][18].

2. POCT of miRNAs

Point-of-care testing (POCT) is defined as testing conducted near or at the site of the patient, and rapid testing may improve patient care [9]. POCT can provide accurate and ultrasensitive disease screening results for patients with the advantages of easy operation, low cost, rapidity, and a visual readout [10][11][12][13]. The development and validation of POCT for early screening of a series of clinical diseases holds great significance. Moreover, POCT provides the possibility of medical guidance and disease screening in remote areas. Recently, POCT has been applied to the rapid and quantitative detection of miRNAs and has made rapid progress. Microfluidics, paper-based biosensors, portable instruments, and visual detection play important roles in POCT and are very promising methods for POCT of miRNAs. To date, dozens of specialized strategies of miRNA detection based on microfluidics and paper-based biosensors have been reported. Microfluidics and paper-based biosensors for miRNA detection have been well reviewed [17][18].

2.1. POCT of miRNAs Based on Portable Instruments

To avoid the need for bulky instruments and auxiliary devices to obtain a high-sensitivity quantitative signal output, we urgently need a sensing strategy that is controllable, low in cost, and independent of sophisticated equipment but that can offer automated readouts for disease-related miRNAs. In this section, the researchers introduce the current situation of the application of off-the-shelf instruments in miRNA detection, analyze and evaluate the possibility and feasibility of their application, and predict their future development trend. A summary of reported POCT methods for miRNAs based on portable instruments is presented below (Table 1).
Table 1. The detection methods of miRNAs based on portable instruments. 

Methods

miRNA

Detection Limit

Samples

Time

Reference

Personal glucose meter

miR-21

0.41 nM/

1 million cells

synthesized miR-21/A549 cell lysates

<2 h

[19]

 

miR-21

10 fM

synthesized miR-21

<2 h

[20]

 

miR-21

miRNA205

2.4 pM

1.1 pM

synthesized miR-21

synthesized miRNA205

<3 h

[21]

 

miR-21

3.65 nM

synthesized miR-21

clinical serum samples from cancer patients

2 h

[22]

 

miR-21

60 pM

3 × 106 cells/mL

synthesized miR-21

MCF-7, A549 and HeLa cell lysates

<3 h

[23]

 

miR-21

68.08 fM

synthesized miR-21

urine samples from DIKI mice

1.5 h

[24]

 

miRNA-155

0.36 fM

synthesized miRNA-155

>5 h

[25]

 

miR-21, miR-335, miR-155, and miR-122

0.325 fmol

synthesized miRNAs

extract from HeLa, HepG2, MCF-7, and L02 Cells

6 h

[26]

Thermometer

miR-21

7.8 nM

synthesized miR-21

HeLa cell lysate

Not mentioned

[27]

 

miRNA-141

0.5 pM

synthesized miRNA-141

>8 h

[28]

Pressure meter

miR-21

7.6 fM

100 cells

synthesized miR-21

A549, MCF-7, HepG2 and HL-7702 cells

20 min

[29]

 

miR-21

10 pM

Serum

0.5 h

[30]

Portable fluorometer

miR-574-5p

2 ng/μL

RNA extract from 5XFAD mice

>3 h

[31]

Capillary force meter

miR-21

10 nM

Human serum

1 h

[32]

 

miR-21

 

MCF-7 cell line

25 min

[33]

Smartphone

miR-133a

0.3 pM

synthesized miR-133a in serum

>5 h

[34]

 

miRNA-499, miRNA-133a

10 fM

synthesized miR-133a in serum

13 h

[35]

 

let-7a

1.7 fM

synthesized let-7a

human serum

2.75 h

[36]

 

miR-133a,

miR-499

1 fM

Synthesized miRNAs

human serum

-

[37]

 

miR-21,

let-7a

fM

Synthesized miRNAs

human serum

<2 h

[38]

 

miR-21

1.43 pM

Synthesized miR-21

human serum, urine

0.5 h

[39]

 

miR-224

1.6 fM

Synthesized miR-224

human plasma

<4.5 h

[40]

 

miR-21

100 fM

500 cells

Synthesized miR-21

MCF-7 and L02 cells

>1 h

[41]

2.2. Visual Detection of miRNAs Based on Colorimetry

Visual detection is particularly attractive for POCT because the readout can be read with the naked eye with no need for instruments. In this section, the researchers summarize recent advances in the visual detection of miRNAs, mainly focusing on colorimetric methods. The researchers provide an all-sided discussion of the principles of the methods and rationally evaluate the applicability of these visual detection methods for early diagnosis based on miRNA detection. A summary of the reported POCT for miRNAs based on colorimetric methods is presented below (Table 2).
Colorimetric assays provide qualitative or quantitative measurement of targets by measuring color changes with no need for special instruments. Detecting a change in color can be used to determine the presence or absence of a target or even to determine its amount range. As color changes can be conveniently judged by the naked eye, colorimetric assays have attracted increasing attention for POCT for miRNAs [12].
Table 2. The detection methods of miRNAs based on visual methods.

Methods

miRNA

Detection Limit

Samples

Time

Reference

Colorimetric detection based on Au-NPs

miR-21

miR-155

5 ng μL−1

Plasma

<3 min

[42]

 

miR-93

miR-223

-

Human serum

-

[43]

 

miR-34a

miR-210

50 ng μL−1

Urine

<20 min

[44]

 

miR-195

40 fM

Human serum

10 min

[45]

 

miR-210-3p

10 pM

Urine

20 min

[46]

 

miR-21

miR-155

1 ng μL−1

Multiple cancerous cell lines and primary fibroblast

<10 min

[47]

 

miR-21

miR-141

3 pM

Synthesized miRNA human serum samples

<5 h

[48]

 

miR-137

0.5 nM

Plasma

1 min

[49]

 

miR-146a

5 nM

Raw cow milk

20 min

[50]

 

let-7a

0.13 pM

A549 cells

50 min

[51]

 

miR-148a

1.9 nM

Plasma

5 min

[52]

 

miR-122

16 pM

Cancerous cell lines

2 h

[53]

 

let-7a

3.13 fM

Human serum

1 h

[54]

 

miR-203

10 pM

MCF-7 cells

-

[55]

 

miR-21

0.23 fM

HeLa, MCF-7, AGS cells

0.5 h

[56]

 

let-7a

4.176 aM

Synthesized let-7a

1 h

[57]

 

miR-221-3p

46 fM

BEL-7404, MDA-MB231, HeLa, and 22Rv1cells

1 h

[58]

 

miR-143

1 fM

Synthesized miR-143

Prostate cancer cell lines VCaP, LNCaP, Du145, and PC-3

>1.5 h

[59]

Colorimetric detection based on enzymatic chromogenic reactions

let-7a

7.4 fM

Synthesized let-7a

2.5 h

[60]

 

miR-122

0.15 aM

Serum

5 min

[61]

 

miR-21

0.2 pM

Serum

50 min

[62]

 

miR-21

1 aM

Serum

<4 h

[63]

 

Let-7a

34 fM

A549 cells

4 h

[64]

 

miR-10b

1 fM

Serum and cell extracts

20 min

[65]

 

miR-141

0.48 nM

Serum

>3 h

[66]

 

miR-21

1 pM

Serum

150 min

[67]

 

miR-141

0.5 pM

Prostate cancer cells

210 min

[68]

 

miR-21

90.3 fM

Serum

<1.5 h

[69]

 

miR-21,

miR-17

1.7 fM

MCF-7

80 min

[70]

 

let-7a

0.1 nM

Serum

3 min

[71]

 

miR-21

44.76 fM

Exosome

2 h

[72]

 

miR-21,

miR-155

0.38 nM

Blood

>1 h

[73]

 

miR-21

4.5 nM

MCF-7 and serum

130 min

[74]

 

miR-21

5 fM

Plasma sample

Cancer cells

Tumor tissues

>6.5 h

[75]

 

miR-155

0.6 pM

Plasma

15 min

[76]

 

miR-205,

miR-944

36.4 fM

Serum

>2 h

[77]

 

miR-155

31.8 fM

Serum

1 h

[78]

 

miR-223

miR-143

20 pM

Synthesized miR-223

iPSCs and CMs

3.5 h

[79]

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