Mrna Detection
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2021 ◽  
Vol 19 (1) ◽  
Hongxia Li ◽  
Antony R. Warden ◽  
Wenqiong Su ◽  
Jie He ◽  
Xiao Zhi ◽  

AbstractPancreatic cancer, at unresectable advanced stages, presents poor prognoses, which could be prevented by early pancreatic cancer diagnosis methods. Recently, a promising early-stage pancreatic cancer biomarker, extracellular vesicles (EVs) related glypican-1 (GPC1) mRNA, is found to overexpress in pancreatic cancer cells. Current mRNA detection methods usually require expensive machinery, strict preservation environments, and time-consuming processes to guarantee detection sensitivity, specificity, and stability. Herein, we propose a novel two-step amplification method (CHAGE) via the target triggered Catalytic Hairpin Assembly strategy combined with Gold-Enhanced point-of-care-testing (POCT) technology for sensitive visual detection of pancreatic cancer biomarker. First, utilizing the catalyzed hairpin DNA circuit, low expression of the GPC1 mRNA was changed into amplification product 1 (AP1, a DNA duplex) as the next detection targets of the paper strips. Second, the AP1 was loaded onto a lateral flow assay and captured with the gold signal nanoparticles to visualize results. Finally, the detected results can be further enhanced by depositing gold to re-enlarge the sizes of gold nanoparticles in detection zones. As a result, the CHAGE methodology lowers the detection limit of mRNA to 100 fM and provides results within 2 h at 37 °C. Furthermore, we demonstrate the successful application in discriminating pancreatic cancer cells by analyzing EVs’ GPC1 mRNA expression levels. Hence, the CHAGE methodology proposed here provides a rapid and convenient POCT platform for sensitive detection of mRNAs through unique probes designs (COVID, HPV, etc.).

2021 ◽  
Vol 2 (3) ◽  
pp. 100711
Giovanni H. Diaz ◽  
Stefan Heller

2021 ◽  
pp. 113507
Dongdong Liu ◽  
Wenhua Li ◽  
Mingzhu Yang ◽  
Lizhen Qiu ◽  
Hongru Pian ◽  

2021 ◽  
Vol 36 (Supplement_1) ◽  
Songtao Feng ◽  
Yueming Gao ◽  
Jingyuan Cao ◽  
Di Yin ◽  
Linli Lv ◽  

Abstract Background and Aims In recent years, it has been found that targeted mRNA detection of sediment cells in urine can be used as novel biomarkers for the diagnosis of diabetic nephropathy (DN). However, its value in predicting the progression of DN is not clear. The purpose of this study is to seek for urinary mRNA markers that evaluate the prognosis of DN through systems biological screening, clinical verification and prospective studies. Method GEO database and “Nephroseq” platform were searched, and the transcriptome data of DN glomeruli and tubules and their clinical information were obtained. Weighted gene co-expression network analysis (WGCNA) combined with gene ontology (GO) annotation and KEGG pathway enrichment analysis were used to screen the hub genes negatively related to eGFR, and the hub genes were used as candidate markers for mRNA detection in urine sediment in DN patients. A total of 91 patients with DN diagnosed by renal biopsy were included, and 60 patients with type 2 diabetes and 61 healthy people were selected as the control groups. The mRNA expression of candidate molecules was detected by Taqman probe quantitative PCR, and the correlation between mRNA expression and eGFR and urinary protein levels were analyzed. Patients with DN were followed up for a median time of 21 months, and the primary end point was defied as end stage renal disease or eGFR decreasing by more than 50%. Multivariate Cox regression was used to evaluate the value of mRNA in predicting DN progression. Results GSE30528 and GSE30529 datasets were selected for analysis, including mRNA expression data of 9 cases of DN and 13 cases of normal glomeruli; and 10 cases of DN and 12 cases of normal tubules respectively. The clinical data of the patients in this study, including gender, race, age and eGFR, were searched on the Nephroseq platform. The gene modules negatively related to eGFR were screened by WGCNA. GO and KEGG analysis showed that the main function of the gene modules in both datasets were related to the activation of inflammatory cells and chemokines pathway. Through the screening of hub genes and the comparison of expression levels, CCL5, CXCL1, CXCL6 and CXCL12 were finally obtained as candidate genes. Quantitative PCR showed that the levels of CCL5 and CXCL1 was significantly increased in DN group, CCL5 was negatively correlated with eGFR and positively correlated with urinary protein level, while CXCL1 was negatively correlated with eGFR, but had no significant correlation with urinary protein level. Multivariate Cox regression showed that eGFR, urinary protein level, degree of renal fibrosis and urinary CCL5 were independent risk factors of primary end point. Conclusion The activation of chemokine signal pathway in renal tissue is involved in the progression of DN. Urinary CCL5 mRNA can independently predict the prognosis of DN and may be served as a novel biomarker for the progression of DN.

2021 ◽  
Yang Li ◽  
Long Mao ◽  
Zongwei Xiao ◽  
Sandeep Bhushan

Abstract BackgroundTo explore whether there is a difference in the expression of ACE and ACE2 genes in patients with acute AD and CHD. MethodsBlood samples from 68 patients, including 34 cases of acute AD (including Stanford type A and B), 21 cases of CHD, and 13 cases of control group. 2 ml of venous blood is submitted for plasma ACE concentration. The arterial wall tissue was taken during the operation for mRNA detection. ResultsThe ACE concentration in the AD group was (17.9 ± 7. 9) U / L, in the CHD group was (33.5 ± 8.1) U / L, and the ACE concentration in the control group was (38.4) ±4.8) U/L, statistically significant (P<0.05). The expression of ACE gene in the AD group was (0.2265 ± 0.3783); the expression in the CHD group was (7.085 7 ± 7.692 9), with significant (P < 0. 05). The expression of ACE2 gene in the AD group was (0.766 2 ± 0.858 6); in the CHD group was (9.612 7 ± 11.542 6), and the difference was significant (P < 0. 05). The expression of the ratio of ACE / ACE2 in the AD group was (0.413 8 ± 0.448); the expression in the CHD group was (0.811 1 ± 0.256 3), the difference was statistically significant (P <0. 001). ConclusionPlasma ACE concentration, ACE and ACE2 gene expression are significantly reduced in acute AD. The imbalance of ACE and ACE2 expression may be involved in the pathogenesis of AD.

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