Multiplexed Detection of Attomoles of Nucleic Acids Using Fluorescent Nanoparticle Counting Platform

Abstract

The sensitive multiplexed detection of nucleic acids in a single sample by a simple manner is of pivotal importance for the diagnosis and therapy of human diseases. Herein, we constructed an automatic fluorescent nanoparticle (FNP) counting platform with a common fluorescence microscopic imaging setup for nonamplification multiplexed detection of attomoles of nucleic acids. Taking the advantages of the highly bright, multicolor emitting FNPs and magnetic separation, the platform enables sensitive multiplexed detection without the need for extra fluorescent labels. Quantification for multiplex DNAs, multiplex microRNAs (miRNA), as well as a DNA and miRNA mixture was achieved with a similar dynamic range, a limit of detection down to 5 amol (5 μL detection volume), and a 81-115% spike recovery from different biological sample matrices. In particular, the sensitivity for multiplex miRNA is by far among the highest without using amplification or the lock nucleic acid hybridization enhancement strategy. Results regarding miRNA-141 from four different cell lines were agreeable with those of the quantitative reverse transcription polymerase chain reaction. Simultaneous detection of miRNA-141 and miRNA-21 in four different cell lines yielded consistent results with publications, indicating the potential for monitoring multiplex miRNA expression associated with the collaborative regulation of important cellular events. This work expands the rule set of multiplex nucleic acid detection strategies and shows promising potential application in clinical diagnosis.

FMSP-Nanoparticles Induced Cell Death on Human Breast Adenocarcinoma Cell Line (MCF-7 Cells): Morphometric Analysis

“…Recently, several attempts have been made to study the effect of different classes of nanoparticles on cancer cells [ 15 , 16 , 17 , 18 , 19 , 20 , 21 , 22 , 23 , 24 , 25 , 26 ]. (Interestingly, fluorescent magnetic nanoparticles have been used in a wide range of applications in biological systems such as diagnostic, bioimaging, and drug delivery [ 27 , 28 , 29 , 30 , 31 , 32 , 33 , 34 , 35 , 36 , 37 , 38 , 39 , 40 , 41 , 42 , 43 , 44 , 45 ] and also in the detection of foodborne pathogens [ 46 ]), but there is no report of use of fluorescent magnetic submicronic polymer nanoparticles (FMSP-nanoparticles) in the treatment of human breast cancer cells. In the present study, we tested the effect of FMSP-nanoparticles on human breast cancer MCF-7 cells.…”

Optical Sensors Based on II-VI Quantum Dots

“…The sensors that use DNA molecules are a great tool not only to detect individual DNA or RNA molecules but also molecules belonging to other classes of biomolecules [83,84,85,86]. Nucleic acids do not have properties that would be useful for their direct detection, so their detection requires the use of, e.g., fluorescent markers [14,87]. Table 5 shows an exemplary QDs-based sensors for nucleic acids determination.…”

“…A complex which was formed between mitoxantrone and ribonucleic acid, prevented more interactions between quantum dots and anticancer drug resulted in enhancing of fluorescence intensity. Quantitative results were obtained for all combinations with a linear range of 20–10,000 pM and a LOD of 3–52 pM [87].…”

Scanning single-molecule counting system for Eprobe with highly simple and effective approach

“…Ultrasensitive detection has also been realized by flow channels [19][20][21][22][23] and rotating containers [24]. To increase the sensitivity, the counting of high-brightness nanoparticles [25,26] and oligonucleotide arrays [27] constructed from the target nucleic acids have been reported. These methods can be used to detect molecules modified with multiple fluorophores or large particles, but they experience difficulty attaining sensitivity beyond the femtomolar level for a single fluorescent molecule.…”