High spatial resolution imaging mass spectrometry and classical histology on a single tissue section

The work presented in this report describes and demonstrates a protocol for protein imaging analysis of biological tissue using MALDI IMS where histological staining and MS analysis are performed on the same tissue section. Spatial image resolution is shown at 35 μm for sagittal sections of the cerebellum from rat brain.     

DNA reorientation on Au nanoparticles: label-free detection of hybridization by surface enhanced Raman spectroscopy

DNA sequences attached to Au nanoparticles via thiol linkers stand up from the surface, giving preferential enhancement of the adenine ring breathing SERS band. Non-specific binding via the nucleobases reorients the DNA, reducing this effect. This change in intensity on reorientation was utilised for label-free detection of hybridization of a molecular beacon.     

Cell processing on polyimide surface patterned by rubbing

In this study, we prepared a novel rubbed fluorinated polyimide film using a rubbing machine with a rubbing cloth and determined the surface properties of the rubbed film using an atomic force microscope and contact angle measurements. In addition, we evaluated the cell adhesion behavior on the rubbed polyimide film using a phase contrast microscope. Interestingly, a rubbed polyimide surface having a micrometer‐scale grooved pattern was prepared by the rubbing method, and the morphologies of rat primary hepatocytes and human liver cell lines attached to the rubbed surface were three‐dimensional multicellular spheroids, while the cells on an unrubbed surface showed two‐dimensional monolayers. This initial study indicates that the rubbing method without any chemical modification is simple and can easily produce large surface areas, suggesting that the rubbing may become a novel cell culture method. Copyright © 2008 John Wiley & Sons, Ltd.     

Direct electrical detection of antigen-antibody binding on diamond and silicon substrates using electrical impedance spectroscopy

The integration of biological molecules with semiconducting materials such as silicon and diamond has great potential for the development of new types of bioelectronic devices, such as biosensors and bioactuators. We have investigated the electrical properties of the antibody-antigen modified diamond and silicon surfaces using electrical impedance spectroscopy (EIS). Frequency dependent measurements at the open-circuit potential show: (a) significant changes in impedance at frequency >10(4) Hz when the surface immobilized IgG was exposed to anti-IgG, and (b) only little or no change when exposed to anti-IgM. Mott-Schottky measurements at high frequency (200 kHz) show that the impedance is dominated by the space charge layer of the semiconducting substrates. Silicon surfaces modified in a similar manner to the diamond surface are compared; n-type and p-type samples show complementary behavior, as expected for a field effect. We also show it is possible to directly observe antigen-antibody interaction at a fixed frequency in real time, and with no additional labeling.     

High mass and spatial resolution mass spectrometry imaging of Nicolas Poussin painting cross section by cluster TOF‐SIMS

Time‐of‐flight secondary ion mass spectrometry (TOF‐SIMS) imaging using cluster primary ion beams is used for the identification of the pigments in the painting of Rebecca and Eliezer at the Well by Nicolas Poussin. The combination of the high mass resolution of the technique with a sub‐micrometer spatial resolution offered by a delayed extraction of the secondary ions, together with the possibility to simultaneously identifying both minerals and organics, has proved to be the method of choice for the study of the stratigraphy of a paint cross section. The chemical compositions of small grains are shown with the help of a thorough processing of the data, with images of specific ions, mass spectra extracted from small regions of interest, and profiles drawn along the different painting layers. Copyright © 2016 John Wiley & Sons, Ltd.     

High sensitive and label-free colorimetric DNA detection based on nicking endonuclease-assisted activation of DNAzymes

Horseradish peroxidase mimicking DNAzyme (HRP-DNAzyme) attracts growing interest as an amplifying label for biorecognition and biosensing events, especially for DNA detection. However, in the traditional designs, one target molecule can only generate one HRP-DNAzyme, which limits the signal enhancement and thus its sensitivity. In this article, we propose an amplified and label-free colorimetric DNA detection strategy based on nicking endonuclease (NEase)-assisted activation of HRP-DNAzymes (NEAA-DNAzymes). This new strategy relies on the hairpin-DNAzyme probe and NEase-assisted target recycling. In the hairpin-DNAzyme probe, the HRP-DNAzyme sequence is protected in a “caged” inactive structure, whereas the loop region includes the target complementary sequence. Upon hybridization with target, the beacon is opened, resulting in the activation of the HRP-DNAzyme. Meanwhile, upon formation of the duplex, the NEase recognizes a specific nucleotide sequence and cleaves the hairpin-DNAzyme probe into two fragments. After nicking, the fragments of the hairpin-DNAzyme probe spontaneously dissociate from the target DNA. Amplification is accomplished by another hairpin-DNAzyme probe hybridizing to the released intact target to continue the strand-scission cycle, which results in activation of numerous DNAzymes. The activated HRP-DNAzymes generate colorimetric or chemiluminescence readout signals, thus providing the amplified detection of DNA. The detection limit of the colorimetric method is 10 pmol/L, which are three orders of magnitude lower than that without NEase. In addition, the detection limit of the chemiluminescence method is 0.2 pmol/L. Meanwhile, this strategy also exhibits high discrimination ability even against single-base mismatch.     

Interfacial electron transfer energetics studied by high spatial resolution tip-enhanced Raman spectroscopic imaging

Interfacial electron transfer (ET) in TiO?-based systems is important in artificial solar energy harvesting systems, catalysis, and in advanced oxidative waste water treatment. The fundamental importance of ET processes and impending applications make the study of interfacial ET a promising research area. Photoexcitation of dye molecules adsorbed on the surface of wide band gap semiconductors, such as TiO?, results in the injection of electrons from the dye molecules to the conduction band of the semiconductor or energetically accessible surface electronic states. Using Raman spectroscopy and ensemble-averaging approaches,t he chemical bonding and vibrational relaxation of the ET processes have been extensively studied. However, due to the complexity of the interfacial ET energetics and dynamics, significant questions remain on characterizing the source of the observed complexities. To address these important issues, we have applied advanced spectroscopic and imaging techniques such as confocal and tip-enhanced near-field Raman as well as photoluminescence spectroscopic and topographic imaging. Here we explore single surface states on TiO? as well as the interfacial electronic coupling of alizarin to TiO? single crystalline surfaces.     

Time of acquisition and high spatial resolution mass spectrometry imaging

The field of mass spectrometry imaging (MSI) is constantly evolving to analyze a diverse array of biological systems. A common goal is the need to resolve cellular and subcellular heterogeneity with high spatial resolution. As the field continues to progress towards high spatial resolution, other parameters must be considered when developing a practical method. Here, we discuss the impacts of high spatial resolution on the time of acquisition and the associated implications they have on an MSI analysis (e.g., area of the region of interest). This work presents a brief tutorial serving to evaluate high spatial resolution MSI relative to time of acquisition and data file size.     

Total internal reflection ellipsometry as a label-free assessment method for optimization of the reactive surface of bioassay devices based on a functionalized cycloolefin polymer

We report a label-free optical detection technique, called total internal reflection ellipsometry (TIRE), which can be applied to study the interactions between biomolecules and a functionalized polymer surface. Zeonor (ZR), a cycloolefin polymer with low autofluorescence, high optical transmittance and excellent chemical resistance, is a highly suitable material for optical biosensor platforms owing to the ease of fabrication. It can also be modified with a range of reactive chemical groups for surface functionalization. We demonstrate the applications of TIRE in monitoring DNA hybridization assays and human chorionic gonadotrophin sandwich immunoassays on the ZR surface functionalized with carboxyl groups. The Ψ and Δ spectra obtained after the binding of each layer of analyte have been fitted to a four-layer ellipsometric model to quantitatively determine the amount of analytes bound specifically to the functionalized ZR surface. Our proposed TIRE technique with its very low analyte consumption and its microfluidic array format could be a useful tool for evaluating several crucial parameters in immunoassays, DNA interactions, adsorption of biomolecules to solid surfaces, or assessment of the reactivity of a functionalized polymer surface towards a specific analyte.     

High spatial resolution in laser-induced breakdown spectroscopy of expanding plasmas

We report a technique that is able to achieve high spatial resolution in the measurement of the temporal and spectral emission characteristics of laser-induced expanding plasmas. The plasma is imaged directly onto the slit of an imaging spectrograph coupled to a time-gated intensified camera, with the plasma expansion direction being parallel to the slit extension. In this way, a single hybrid detection system is used to acquire the spatial, spectral and temporal characteristics of the laser induced plasma. The parallel acquisition approach of this technique ensures a much better spatial resolution in the expansion direction, reproducibility and data acquisition speed than commonly obtained by sequential measurements at different distances from the target. We have applied this technique to study the laser-induced plasma in LiNbO₃ and Bi₁₂Ge₁O₂₀, revealing phenomena not seen in such detail with standard instruments. These include extreme line broadening up to a few nanometers accompanied by self-absorption near the target surface, as well as different ablation and expansion dynamics for the different species ejected. Overall, the high precision and wealth of quantitative information accessible with this technique open up new possibilities for the study of fundamental plasma expansion processes during pulsed laser ablation.     

High resolution slice imaging of a molecular speed distribution

High resolution slice imaging experiments are reported measuring the speed distribution of molecular fragments, recoiling at a most probable speed v(mp), with a full-width-half-maximum (FWHM) speed resolution near the permille level: FWHM(v)/v(mp) = 1.9 x 10(-3). We implemented a high resolution single-particle slice imaging detector and used a two-colour resonance-enhanced multi-photon ionisation (REMPI) scheme to reduce broadening of the speed distribution due to the electron kick. The results on the measurement of the CD(3) speed distribution from photolysis of CD(3)I show that it is possible to image the three-dimensional speed distribution at a resolution down to FWHM(v) = 6.7 m s(-1), when the fragment has an absolute speed of v(mp) = 3473 m s(-1). The new experiments demonstrate the potential of slice imaging to measure with high resolution the three-dimensional speed distribution of a cloud of molecules.     

High resolution depth profile analysis by elastic recoil detection with heavy ions

Elastic recoil detection (ERD) with energetic heavy ions (e.g. 60-120 MeV(127)I) is a suitable method to measure depth profiles of light and medium heavy elements in thin films. The advantages of this method are reliable and quantitative results and elementally and isotopically resolved depth profiles. A relative energy resolution of 0.07% has been measured in real ERD-experiments using the Q3D magnetic spectrograph at the Munich tandem accelerator and a large solid angle of detection of 5 msr. The good energy resolution allows atomic depth resolution near to the surface which has been obtained at flat and smooth carbon samples. A large solid angle of detection is necessary to measure a depth profile with the desired accuracy before the sample is significantly altered by the ion beam. As an example carbon profiles of thin carbon layers, prepared by a laser plasma ablation deposition process, have been investigated revealing the high depth resolution and its power to resolve elemental profiles at gradiated interfaces.     

High resolution depth profile analysis by elastic recoil detection with heavy ions

Elastic recoil detection (ERD) with energetic heavy ions (e.g. 60–120 MeV¹²⁷I) is a suitable method to measure depth profiles of light and medium heavy elements in thin films. The advantages of this method are reliable and quantitative results and elementally and isotopically resolved depth profiles. A relative energy resolution of 0.07% has been measured in real ERD-experiments using the Q3D magnetic spectrograph at the Munich tandem accelerator and a large solid angle of detection of 5 msr. The good energy resolution allows atomic depth resolution near to the surface which has been obtained at flat and smooth carbon samples. A large solid angle of detection is necessary to measure a depth profile with the desired accuracy before the sample is significantly altered by the ion beam. As an example carbon profiles of thin carbon layers, prepared by a laser plasma ablation deposition process, have been investigated revealing the high depth resolution and its power to resolve elemental profiles at gradiated interfaces.     

Aptamer biosensor for label-free impedance spectroscopy detection of proteins based on recognition-induced switching of the surface charge

The recognition of proteins by aptamer-modified electrode transducers reverses the surface charge and leads to a novel label-free impedance spectroscopy bioelectronic detection protocol based on a decrease in the electron transfer resistance.     

Phase transition of individually addressable microstructured membranes visualized by imaging ellipsometry

The phase transition of individually addressable microstructured lipid bilayers was investigated by means of imaging ellipsometry. Microstructured bilayers were created on silicon substrates by micromolding in capillaries, and the thermotropic behavior of various saturated diacyl phosphatidylcholine (1,2-dipalmitoyl-sn-glycero-3-phosphocholine, 1,2-dipentadecoyl-sn-glycero-3-phosphocholine, and 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC)) bilayers as well as DMPC/cholesterol membranes was determined by measuring the area expansion and thickness of the bilayer as a function of temperature. We found an increase in the main phase transition temperature T(M) of 2-6 degrees C and a substantially reduced cooperativity compared to multilamellar vesicles. Measurements of lateral diffusion constants D employing fluorescence recovery after photobleaching revealed, however, only a marginal decrease in D compared to those found for vesicles and multibilayers. The known dependencies of T(M) both on the chain length of diacyl PC membranes and on the cholesterol content were reproduced on a solid support. Microstructured bilayers offer the unique advantage of integrating an internal standard of known thermotropic properties, which turned out to be important for reducing the measurement error and for ruling out the slightly changing impact of the surface on the phase transition behavior due to the surface pretreatment.     

High resolution imaging of functional group distributions on carbon surfaces by tapping mode atomic force microscopy

Here we report, for the first time, the high resolution imaging of hydrophilic, polar functional group distributions on flat carbon surfaces by phase contrast in noncontact tapping mode AFM.