Aptamer–nanoparticle complexes as powerful diagnostic and therapeutic tools

Correct diagnosis and successful therapy are extremely important to enjoy a healthy life when suffering from a disease. To achieve these aims, various cutting-edge technologies have been designed and fabricated to diagnose and treat specific diseases. Among these technologies, aptamer–nanomaterial hybrids have received considerable attention from scientists and doctors because they have numerous advantages over other methods, such as good biocompatibility, low immunogenicity and controllable selectivity. In particular, aptamers, oligonucleic acids or peptides that bind to a specific target molecule, are regarded as outstanding biomolecules. In this review, several screening techniques for aptamers, also called systematic evolution of ligands by exponential enrichment (SELEX) methods, are introduced, and diagnostic and therapeutic aptamer applications are also presented. Furthermore, we describe diverse aptamer–nanomaterial conjugate designs and their applications for diagnosis and therapy.112618Ysciescopuskc

Crystal structure of a DNA aptamer bound to PvLDH elucidates novel single-stranded DNA structural elements for folding and recognition

Structural elements are key elements for understanding single-stranded nucleic acid folding. Although various RNA structural elements have been documented, structural elements of single-stranded DNA (ssDNA) have rarely been reported. Herein, we determined a crystal structure of PvLDH in complex with a DNA aptamer called pL1. This aptamer folds into a hairpin-bulge contact by adopting three novel structural elements, viz, DNA T-loop-like motif, base-phosphate zipper, and DNA G.G metal ion zipper. Moreover, the pL1: PvLDH complex shows unique properties compared with other protein: nucleic acid complexes. Generally, extensive intermolecular hydrogen bonds occur between unpaired nucleotides and proteins for specific recognitions. Although most protein-interacting nucleotides of pL1 are unpaired nucleotides, pL1 recognizes PvLDH by predominant shape complementarity with many bridging water molecules owing to the combination of three novel structural elements making protein-binding unpaired nucleotides stable. Moreover, the additional set of Plasmodium LDH residues which were shown to form extensive hydrogen bonds with unpaired nucleotides of 2008s does not participate in the recognition of pL1. Superimposition of the pL1: PvLDH complex with hLDH reveals steric clashes between pL1 and hLDH in contrast with no steric clashes between 2008s and hLDH. Therefore, specific protein recognition mode of pL1 is totally different from that of 2008s.1152Ysciescopu

Formation of lipid bilayer membrane in a poly(dimethylsiloxane) microchip integrated with a stacked polycarbonate membrane support and an on-site nanoinjector

This paper describes a new and facile approach for the formation of pore-spanning bilayer lipid membranes (BLMs) within a poly(dimethylsiloxane) (PDMS) microfluidic device. Commercially, readily available polycarbonate (PC) membranes are employed for the support of BLMs. PC sheets with 5 mu m, 2 mu m, and 0.4 mu m pore diameters, respectively, are thermally bonded into a multilayer-stack, reducing the pore density of 0.4 mu m-pore PC by a factor of 200. The BLMs on this support are considerably stable (a mean lifetime: 17 h). This multilayer-stack PC (MSPC) membrane is integrated into the PDMS chip by an epoxy bonding method developed to secure durable bonding under the use of organic solvents. The microchip has a special channel for guiding a micropipette in the proximity of the MSPC support. With this on-site injection technique, tens to hundreds of nanoliters of solutions can be directly dispensed to the support. Incorporating gramicidin ion channels into BLMs on the MSPC support has confirmed the formation of single BLMs, which is based on the observation from current signals of 20 pS conductance that is typical to single channel opening. Based on the bilayer capacitance (1.4 pF), about 15% of through pores across the MSPC membrane are estimated to be covered with BLMs. (C) 2015 AIP Publishing LLC.open110sciescopu

Aptamers and Their Biological Applications

Recently, aptamers have attracted the attention of many scientists, because they not only have all of the advantages of antibodies, but also have unique merits, such as thermal stability, low cost, and unlimited applications. In this review, we present the reasons why aptamers are known as alternatives to antibodies. Furthermore, several types of in vitro selection processes, including nitrocellulose membrane filtration, affinity chromatography, magnetic bead, and capillary electrophoresis-based selection methods, are explained in detail. We also introduce various applications of aptamers for the diagnosis of diseases and detection of small molecules. Numerous analytical techniques, such as electrochemical, colorimetric, optical, and mass-sensitive methods, can be utilized to detect targets, due to convenient modifications and the stability of aptamers. Finally, several medical and analytical applications of aptamers are presented. In summary, aptamers are promising materials for diverse areas, not just as alternatives to antibodies, but as the core components of medical and analytical equipment

Electrochemical detection of mismatched DNA using a MutS probe

A direct and label-free electrochemical biosensor for the detection of the protein–mismatched DNA interaction was designed using immobilized N-terminal histidine tagged Escherichia coli. MutS on a Ni-NTA coated Au electrode. General electrochemical methods, cyclic voltammetry (CV), electrochemical quartz crystal microbalance (EQCM) and impedance spectroscopy, were used to ascertain the binding affinity of mismatched DNAs to the MutS probe. The direct results of CV and impedance clearly reveal that the interaction of MutS with the CC heteroduplex was much stronger than that with AT homoduplex, which was not differentiated in previous results (GT > CT > CC ≈ AT) of a gel mobility shift assay. The EQCM technique was also able to quantitatively analyze MutS affinity to heteroduplexes

Simultaneous electrochemical detection of both PSMA (+) and PSMA (-) prostate cancer cells using an RNA/peptide dual-aptamer probe

Using an RNA/peptide dual-aptamer probe, both PSMA (+) and PSMA (-) prostate cancer cells were simultaneously detected by electrochemical impedance spectroscopy. This approach can be applied as a general tool for early diagnosis of prostate cancer.CATALONA WJ, 1993, JAMA-J AM MED ASSOC, V270, P948Lupold SE, 2002, CANCER RES, V62, P4029Kue PF, 2002, INT J CANCER, V102, P572, DOI 10.1002/ijc.10734Drummond TG, 2003, NAT BIOTECHNOL, V21, P1192, DOI 10.1038/nbt873DARAIN F, 2004, BIOSENS BIOELECTRON, V20, P856Ban CG, 2004, NUCLEIC ACIDS RES, V32, DOI 10.1093/nar/gnh109Ghosh A, 2004, J CELL BIOCHEM, V91, P528, DOI 10.1002/jcb.10661LEVIN MA, 2005, J UROLOGY, V159, P475Rodriguez MC, 2005, CHEM COMMUN, P4267, DOI 10.1039/b506571bZitzmann S, 2005, CLIN CANCER RES, V11, P139Horninger W, 2001, CANCER-AM CANCER SOC, V91, P1667Lang SH, 2001, BRIT J CANCER, V85, P590Yamamoto T, 2001, UROLOGY, V58, P994Palecek E, 2002, CRIT REV ANAL CHEM, V32, P261Narain V, 2002, CANCER METAST REV, V21, P17Edwards S, 2005, BRIT J CANCER, V92, P376, DOI 10.1038/sj.bjc.6602261Postma R, 2005, EUR J CANCER, V41, P825, DOI 10.1016/j.ejca.2004.12.029Cahova-Kucharikova K, 2005, ANAL CHEM, V77, P2920Rahman MA, 2005, ANAL CHEM, V77, P4854, DOI 10.1021/ac050558vCho M, 2006, NUCLEIC ACIDS RES, V34, DOI 10.1093/nar/gkl364Farokhzad OC, 2006, P NATL ACAD SCI USA, V103, P6315, DOI 10.1073/pnas.0601755103Chu TC, 2006, CANCER RES, V66, P5989, DOI 10.1158/0008-5472.CAN-05-4583McNamara JO, 2006, NAT BIOTECHNOL, V24, P1005, DOI 10.1038/nbt1223Palecek E, 1998, BIOSENS BIOELECTRON, V13, P621Min K, 2008, BIOSENS BIOELECTRON, V23, P1819, DOI 10.1016/j.bios.2008.02.021CHO M, 2008, BMB REPORTS, V41, P119Kim D, 2007, J AM CHEM SOC, V129, P7661, DOI 10.1021/ja071471pMaalouf R, 2007, ANAL CHEM, V79, P4879, DOI 10.1021/ac070085nKRAHN MD, 1994, JAMA-J AM MED ASSOC, V272, P773

Cardioprotective Effect of the SDF-1α/CXCR4 Axis in Ischemic Postconditioning in Isolated Rat Hearts

Background and Objectives: Information about the role of the stromal cell-derived factor1a (SDF-1 alpha)/chemokine receptor type 4 (CXCR4) axis in ischemic postconditioning (IPOC) is currently limited. We hypothesized that the SDF-1 alpha/CXCR4 signaling pathway is directly involved in the cardioprotective effect of IPOC. Methods: Isolated rat hearts were divided into four groups. The control group was subjected to 30-min of regional ischemia and 2-hour of reperfusion (n=12). The IPOC group was induced with 6 cycles of 10-second reperfusion and 10-second global ischemia (n=8) in each cycle. The CXCR4 antagonist, AMD3100, was applied before reperfusion in the IPOC group (AMD+IPOC group, n=11) and control group (AMD group, n=9). Hemodynamic changes with electrocardiography were monitored and infarct size was measured. The SDF-1 alpha, lactate dehydrogenase (LDH) and creatine kinase (CK) concentrations in perfusate were measured. We also analyzed extracellular signal-regulated kinase 1/2 (ERK1/2) and Akt phosphorylation state expression. Results: IPOC significantly reduced infarct size, but AMD3100 attenuated the infarct reducing effect of IPOC. IPOC significantly decreased LDH and CK, but these effects were reversed by AMD3100. ERK1/2 and Akt phosphorylation increased with IPOC and these effects were blocked by AMD3100. Conclusion: Based on the results of this study, SDF-1 alpha/CXCR4 signaling may be involved in IPOC cardioprotection and this signaling pathway couples to the ERK1/2 and Akt pathways.111Ysciescopuskc

Cardioprotective Effect of the SDF-1α/CXCR4 Axis in Ischemic Postconditioning in Isolated Rat Hearts

Information about the role of the stromal cell-derived factor-1α (SDF-1α)/chemokine receptor type 4 (CXCR4) axis in ischemic postconditioning (IPOC) is currently limited. We hypothesized that the SDF-1α/CXCR4 signaling pathway is directly involved in the cardioprotective effect of IPOC

Single-Molecule Analysis Reveals the Kinetics and Physiological Relevance of MutL-ssDNA Binding

DNA binding by MutL homologs (MLH/PMS) during mismatch repair (MMR) has been considered based on biochemical and genetic studies. Bulk studies with MutL and its yeast homologs Mlh1-Pms1 have suggested an integral role for a single-stranded DNA (ssDNA) binding activity during MMR. We have developed single-molecule Förster resonance energy transfer (smFRET) and a single-molecule DNA flow-extension assays to examine MutL interaction with ssDNA in real time. The smFRET assay allowed us to observe MutL-ssDNA association and dissociation. We determined that MutL-ssDNA binding required ATP and was the greatest at ionic strength below 25 mM (KD = 29 nM) while it dramatically decreases above 100 mM (KD>2 µM). Single-molecule DNA flow-extension analysis suggests that multiple MutL proteins may bind ssDNA at low ionic strength but this activity does not enhance stability at elevated ionic strengths. These studies are consistent with the conclusion that a stable MutL-ssDNA interaction is unlikely to occur at physiological salt eliminating a number of MMR models. However, the activity may infer some related dynamic DNA transaction process during MMR