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Nanostructured paper-based platform for phenylalanine neonatal screening by LED-induced fluorescence
摘要： In this work, a novel paper-based analytical device (PAD) coupled to LED-induced fluorescence (LIF) detection (fPAD) for the rapid, selective, and sensitive quantification of phenylalanine (Phe) in neonatal samples was developed. Enzymes Phenylalanine dehydrogenase (PheDH) and diaphorase were immobilized on a paper microzone previously modified with zinc oxide nanoparticles (ZnONPs) coated with chitosan (CH-ZnONPs). Phe was extracted from the blood spots collected samples on filter paper and was mixed with nicotinamide adenine dinucleotide (NAD+) and resazurin. Then the mixture was deposited on the reaction microzone of the fPAD where PheDH converts the Phe and NAD+ to phenylpyruvate and NADH, respectively. Finally, NADH was oxidized by diaphorase with the consequent reduction from resazurin to resorufin. This latter was detected by LIF using an excitation wavelength of 535 nm and an emission of 580 nm in a synchronized video microscope. We compare the responses of the PADs with and without nanomaterials to demonstrate the improved analytical performance of the developed devices. For this, the PADs were modified with the same concentration of horseradish peroxidase (HRP). The fluorescent signal obtained from the PADs with nanomaterials was higher than that of the unmodified PADs. Our method exhibited within- and between-assay variation coefficients below 5.23% and 6.67%, respectively. The detection limit obtained by the developed device was 0.125 μM. The proposed fPAD allowed the simple, rapid, low-cost, and sensitive detection of Phe in neonatal blood samples.
关键词： zinc oxide nanoparticles,Phenylalanine,Paper-based analytical device,Enzymatic method,Fluorometric detection
更新于2025-09-23 15:19:57
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Multiplexed determination of intracellular messenger RNA by using a graphene oxide nanoprobe modified with target-recognizing fluorescent oligonucleotides
摘要： A multiplexed graphene oxide (GO) fluorescent nanoprobe is described for quantification and imaging of messenger RNAs (mRNAs) in living cells. The recognizing oligonucleotides (with sequences complementary to those of target the fluorescence of the recognizing mRNAs) were labeled with different fluorescent dyes. If adsorbed on GO, oligonucleotides is quenched. After having penetrated living cells, the oligonucleotides bind to target mRNAs and dissociate from GO. This leads to the recovery of fluorescence. Using different fluorescent dyes, various intracellular mRNAs can be simultaneously imaged and quantified by a high content analysis within a short period of time. Actin mRNA acts as the internal control. This GO-based nanoprobe allows mRNA mimics to be determined within an analytical range from 1 to 400 nM and a detection limit as low as 0.26 nM. Up to 3 intracellular mRNAs (C-myc, TK1, and actin) can be detected simultaneously in a single living cell. Hence, this nanoprobe enables specific distinction of intracellular mRNA expression levels in cancerous and normal cells. It can be potentially applied as a tool for detection of cancer progression and diagnosis.
关键词： Fluorescence resonance energy transfer (FRET),Cancer biomarker,Actin mRNA,Fluorometric detection,High content analysis,Cancer diagnosis
更新于2025-09-10 09:29:36