- A significant enhancement of color transition from an on-off type achromatic colorimetric nanosensor for highly sensitive multi-analyte detection with the naked eye. [Journal Article]
- NNanoscale 2016 Oct 20
- Here, we report the development of an achromatic nanoparticle-based colorimetric sensor (achromatic nanosensor) with an on-off type color change that significantly enhances the color transition and i...
Here, we report the development of an achromatic nanoparticle-based colorimetric sensor (achromatic nanosensor) with an on-off type color change that significantly enhances the color transition and increases the sensitivity of the sensor for naked-eye inspection. The achromatic nanosensor was prepared via a modified CMYK (CRYK) subtractive color model by combining DNA-functionalized gold nanoparticles (AuNPs-DNA), silver nanoparticles (AgNPs-DNA), and gold nanorods (AuNRs-DNA). The initially black-colored achromatic nanosensor not only allowed multiplexed detection by generating target-specific diverse color changes, but also improved the recognition of color changes by the naked eye. Thus, this on-off type color change enabled analysis near the limit of detection (LOD) with the naked eye. In addition, we developed a new image processing method adapted for this achromatic sensor. By quantifying the saturation value of the color images of the achromatic sensor, we could significantly amplify the color signal of the samples, which is difficult to achieve with general colorimetric sensors. The practical application of this achromatic nanosensor for biomarker detection was demonstrated with thrombin and platelet-derived growth factor (PDGF) in human blood plasma. These results provide a new sensing platform that is applicable to most NP-based colorimetric sensing systems for a wide range of applications, including biomolecular diagnosis, chemical pollutant sensing, environmental monitoring, etc.
- Nucleosomes Selectively Inhibit Cas9 Off-Target Activity at a Site Located at the Nucleosome Edge. [Journal Article]
- JBJ Biol Chem 2016 Oct 18
- Nucleosomes affect Cas9 binding and activity at on-target sites, but their impact at off-target sites is unknown. To investigate how nucleosomes affect Cas9 cleavage at off-target sites in vitro, we ...
Nucleosomes affect Cas9 binding and activity at on-target sites, but their impact at off-target sites is unknown. To investigate how nucleosomes affect Cas9 cleavage at off-target sites in vitro, we used a single guide RNA (sgRNA) that has been previously shown to efficiently direct Cas9 cleavage at the edge of the strongly positioned 601 nucleosome. Our data indicate that single mismatches between the sgRNA and DNA target have relatively little effect on Cas9 cleavage of naked DNA substrates, but strongly inhibit cleavage of nucleosome substrates, particularly when the mismatch is in the sgRNA 'seed' region. These findings indicate that nucleosomes may enhance Cas9 specificity by inhibiting cleavage of off-target sites at the nucleosome edge.
- Induction of HIV-1 gag specific immune responses by cationic micelles mediated delivery of gag mRNA. [Journal Article]
- DDDrug Deliv 2016; 23(7):2596-2607
- In recent years, mRNA-based vaccines have emerged to be a great alternative to DNA-based vaccines due to the safety of not inserting into host genome. However, mRNA molecules are single-stranded nucl...
In recent years, mRNA-based vaccines have emerged to be a great alternative to DNA-based vaccines due to the safety of not inserting into host genome. However, mRNA molecules are single-stranded nucleic acids that are vulnerable under RNase existing in human skin and tissues. In this study, a self-assembled cationic nanomicelles based on polyethyleneimine-stearic acid (PSA) copolymer were developed to delivery HIV-1 gag encoding mRNA to dendritic cells and BALB/c mice. We evaluated the transfection efficiency and cell uptake efficiency of naked EGFP mRNA, PSA, PEI-2k and PEI-25k nanoparticles format on DC2.4 cell lines. Immune responses after sub-cutaneous administration of gag mRNA to BALB/c mice were notably induced by PSA as compared with naked gag mRNA. We found the PSA/mRNA nanomicelles were potent systems that can effectively deliver mRNA and induce antigen-specific immune response, stimulating various new vaccine strategies using mRNA.
- A capillary-based multiplexed isothermal nucleic acid-based test for sexually transmitted diseases in patients. [Journal Article]
- CCChem Commun (Camb) 2016 Oct 6; 52(82):12187-12190
- We demonstrate a multiplexed loop mediated isothermal amplification (LAMP) assay for infectious disease diagnostics, where the analytical process flow of target pathogens genomic DNA is performed man...
We demonstrate a multiplexed loop mediated isothermal amplification (LAMP) assay for infectious disease diagnostics, where the analytical process flow of target pathogens genomic DNA is performed manually by moving magnetic beads through a series of plugs in a capillary. Heat is provided by a water bath and the results are read by the naked eye, enabling applications in low resource settings.
- Tissue-specific Calibration of Real-time PCR Facilitates Absolute Quantification of Plasmid DNA in Biodistribution Studies. [Journal Article]
- MTMol Ther Nucleic Acids 2016 Oct 4; 5(10):e371
- Analysis of the tissue distribution of plasmid DNA after administration of nonviral gene delivery systems is best accomplished using quantitative real-time polymerase chain reaction (qPCR), although ...
Analysis of the tissue distribution of plasmid DNA after administration of nonviral gene delivery systems is best accomplished using quantitative real-time polymerase chain reaction (qPCR), although published strategies do not allow determination of the absolute mass of plasmid delivered to different tissues. Generally, data is expressed as the mass of plasmid relative to the mass of genomic DNA (gDNA) in the sample. This strategy is adequate for comparisons of efficiency of delivery to a single site but it does not allow direct comparison of delivery to multiple tissues, as the mass of gDNA extracted per unit mass of each tissue is different. We show here that by constructing qPCR standard curves for each tissue it is possible to determine the dose of intact plasmid remaining in each tissue, which is a more useful parameter when comparing the fates of different formulations of DNA. We exemplify the use of this tissue-specific qPCR method by comparing the delivery of naked DNA, cationic DNA complexes, and neutral PEGylated DNA complexes after intramuscular injection. Generally, larger masses of intact plasmid were present 24 hours after injection of DNA complexes, and neutral complexes resulted in delivery of a larger mass of intact plasmid to the spleen.
- Studying Closed Hydrodynamic Models of "In Vivo" DNA Perfusion in Pig Liver for Gene Therapy Translation to Humans. [Journal Article]
- PlosPLoS One 2016; 11(10):e0163898
- CONCLUSIONS: Hydrofection of hAAT DNA to "in vivo" isolated pig liver mediates highly efficient gene delivery and protein expression in tissue. Both endovascular and surgically closed models mediate high tissue protein expression. Impairment of protein secretion to plasma is observed and might be species-related. This study reinforces the potential application of closed liver hydrofection for therapeutic purposes, provided protein secretion improves.
- Rapid and Visual Chlamydia trachomatis Detection using Loop-Mediated Isothermal Amplification and Hydroxynaphthal Blue. [Journal Article]
- LALett Appl Microbiol 2016 Sep 30
- We developed an assay comprising crude DNA lysis by simple heat treatment coupled loop-mediated isothermal amplification with hydroxynaphthol blue (LAMP-HNB) for C. trachomatis detection, and evaluat...
We developed an assay comprising crude DNA lysis by simple heat treatment coupled loop-mediated isothermal amplification with hydroxynaphthol blue (LAMP-HNB) for C. trachomatis detection, and evaluated the developed assay for its feasibility as one-step point-of-care detection on 284 endocervical swab specimens from clinically symptomatic C. trachomatis and healthy subjects. This assay is sensitive to 0.04 pg of ompA, specific with 6 primers targeting C. trachomatis ompA region, rapid (45 min total assay time), inexpensive (~3 USD/reaction), does not require sophisticated instrumentation, and has comparable assay effectiveness (95% specificity, 90-100% sensitivity) to the bacterial DNA isolation by commercial kit coupled with PCR and gel electrophoresis (98-100% specificity, 87-100% sensitivity) based on the clinical samples test. The test result could be read by naked eyes through the color change from violet (negative) to sky blue (positive) color for C. trachomatis-infected specimen. Further, this assay uses all safe chemical reagents and is hence safe to the users. This article is protected by copyright. All rights reserved.
- Smart Carbon Nanotubes with Laser-Controlled Behavior in Gene Delivery and Therapy through a Non-Digestive Trafficking Pathway. [Journal Article]
- SSmall 2016 Sep 28
- Near-infrared (NIR) laser-controlled gene delivery presents some benefits in gene therapy, inducing enhanced gene transfection efficiency. In this study, a "photothermal transfection" agent is obtain...
Near-infrared (NIR) laser-controlled gene delivery presents some benefits in gene therapy, inducing enhanced gene transfection efficiency. In this study, a "photothermal transfection" agent is obtained by wrapping poly(ethylenimine)-cholesterol derivatives (PEI-Chol) around single-walled carbon nanotubes (SWNTs). The PEI-Chol modified SWNTs (PCS) are effective in compressing DNA molecules and protecting them from DNaseI degradation. Compared to the complexes formed by PEI with DNA (PEI/DNA), complexes of PCS and DNA that are formed (PCS/DNA) exhibit a little lower toxicity to HEK293 and HeLa cells under the same PEI molecule weight and weight ratios. Notably, caveolae-mediated cellular uptake of PCS/DNA occurs, which results in a safer intracellular transport of the gene due to the decreased lysosomal degradation in comparison with that of PEI/DNA whose internalization mainly depends on clathrin rather than caveolae. Furthermore, unlike PEI/DNA, PCS/DNA exhibits a photothermal conversion ability, which promotes DNA release from PCS under NIR laser irradiation. The NIR laser-mediated photothermal transfection of PCS10K /plasmid TP53 (pTP53) results in more apoptosis and necrosis of HeLa cells in vitro than other groups, and achieves a higher tumor-growth inhibition in vivo than naked pTP53, PEI25K /pTP53, and PCS10K /pTP53 alone. The enhanced transfection efficiency of PCS/DNA can be attributed to more efficient DNA internalization into the tumor cells, promotes detachment of DNA from PCS under the mediation of NIR laser and higher DNA stability in the cells due to caveolae-mediated cellular uptake of the complexes.
- Detection of parvovirus B19 DNA in blood: Viruses or DNA remnants? [Journal Article]
- JCJ Clin Virol 2016 Sep 13; 84:19-23
- CONCLUSIONS: Detection of B19V DNA in blood by PCR does not necessarily imply that B19V replication takes place and that infectious B19V virions are present. We propose that remnant B19V DNA strands can be released from tissues without active replication. This finding urges to reconsider an assumed role of B19V infection mainly based on B19V DNA detection in blood, a much debated subject in clinical syndromes such as myocarditis and arthritis.
New Search Next
- Multiplexed instrument-free meningitis diagnosis on a polymer/paper hybrid microfluidic biochip. [Journal Article]
- BBBiosens Bioelectron 2016 Sep 12; 87:865-873
- Neisseria meningitidis (N. meningitidis), Streptococcus pneumoniae (S. pneumoniae), and Haemophilus influenzae type b (Hib) are three most common pathogens accounting for most bacterial meningitis, a...
Neisseria meningitidis (N. meningitidis), Streptococcus pneumoniae (S. pneumoniae), and Haemophilus influenzae type b (Hib) are three most common pathogens accounting for most bacterial meningitis, a serious global infectious disease with high fatality, especially in developing nations. Because the treatment and antibiotics differ among each type, the identification of the exact bacteria causing the disease is vital. Herein, we report a polymer/paper hybrid microfluidic biochip integrated with loop-mediated isothermal amplification (LAMP) for multiplexed instrument-free diagnosis of these three major types of bacterial meningitis, with high sensitivity and specificity. Results can be visually observed by the naked eye or imaged by a smartphone camera under a portable UV light source. Without using any specialized laboratory instrument, the limits of detection of a few DNA copies per LAMP zone for N. meningitidis, S. pneumoniae and Hib were achieved within 1h. In addition, these three types of microorganisms spiked in artificial cerebrospinal fluid (ACSF) were directly detected simultaneously, avoiding cumbersome sample preparation procedures in conventional methods. Compared with the paper-free non-hybrid microfluidic biochip over a period of three months, the hybrid microfluidic biochip was found to have a much longer shelf life. Hence, this rapid, instrument-free and highly sensitive microfluidic approach has great potential for point-of-care (POC) diagnosis of multiple infectious diseases simultaneously, especially in resource-limited settings.