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PubMed2002

Patterning DNA on microm scale on mica.

Double-stranded DNA molecules were patterned by selective adsorption to aminosilane patterns on mica surfaces. Line patterns with 10 microm spacing were made by photolithography and transferred to a polymer stamp. The stamp was then used for applying aminosilane molecules by microcontact-printing technique on mica substrates. We applied DNA in Tris-EDTA (TE) buffer solution on the patterned substrate, and incubated it for 5 min at room temperature. The sample was then rinsed with pure water, and dried with nitrogen gas. Tapping mode force microscopy showed that DNA was adsorbed selectively on the aminosilanized parts of the mica substrate. We also tried to bridge two aluminum electrodes with DNA using AC electrophoresis.

Adsorption↗

PubMed1999

Controlled partial skin thickness burns: an animal model for studies of burnwound progression.

A practicable, reliable and reproducible model for infliction of partial skin thickness burn lesions in rabbits is presented. The model is dedicated to experimental studies investigating the influence of drugs on burn wounds. A round aluminium stamp with a contact area of 4 cm2, weight 85 g, was heated up to 80 degrees C and applied for 14 s without additional pressure on the depilated dorsal skin of rabbits. This procedure produced the desired partial skin thickness burn injury. The depth of the burn lesions was investigated by HE-stained paraffin sections. The border of the necrotic zone was found in the central third of the dermis in 80% of cases, and in the central two quarters in 100%. These results are achieved when the rabbit's hair at the site of infliction is in the anagen phase of the hair growth cycle. For obtaining reproducible results we recommend using rabbits of the same strain and weight, anagen hair growth phase, the described procedure of infliction, an identical stamp and the specified temperature and infliction time.

Animals↗

PubMed2000

Introduction to classical swine fever: virus, disease and control policy.

Classical swine fever virus is a spherical enveloped particle of about 40-60 nm in diameter with a single stranded RNA genome of about 12,300 bases with positive polarity, classified as a pestivirus within the family Flaviviridae. Natural hosts are domestic and wild pigs. The virus causes one of the most severe diseases in pigs world wide with grave economic consequences. The clinical picture of classical swine fever is variable, depending on the age of the affected animals and viral virulence. The virus is well characterised and reliable laboratory diagnostic procedures are available. In many parts of the world live attenuated vaccines are being used as a safe and efficient prophylactic tool. However, in EU Member States and several other countries vaccination is prohibited and CSF is controlled by a strict stamping out policy. In order to overcome the disadvantages of conventional vaccination inactivated marker vaccines have been developed that enable the distinction between vaccinated and infected animals. Whether these vaccines will be accepted as an additional tool in the framework of the stamping out policy is not yet decided.

Animals↗

PubMed1994

A criterion based audit of community-acquired pneumonia.

UNLABELLED: We undertook a prospective audit of patients admitted to a specialist respiratory unit in order to assess: (a) the completeness of the objective assessment of severity of community-acquired pneumonia in patients on admission to hospital, and whether such indices were useful in predicting outcome; (b) the completeness of microbiological testing and whether maximizing the number of specimens sent to the laboratory would increase the frequency of positive microbiological diagnoses and; (c) the influence of the above on clinical management. Forty-eight patients with consolidation on chest X-ray were studied in two audit periods: 1 February 1991 to 1 May 1991 and 2 May 1991 to 16 March 1992. After the first audit period, a rubber stamp with a check list of microbiological investigations was used in the patients' notes. Seven 'markers of severity' of pneumonia, were recorded consistently throughout the study and these were useful in predicting outcome (P < 0.01). In the first audit period sputum culture, blood culture and acute serology were recorded in approximately 50% of patients. Following the introduction of the rubber stamp, the requesting of sputum, blood and urine antigen rose from none in the first audit period to 40-60%. The increase in the number of specimens obtained was not associated with an increase in the proportion of positive microbiological diagnoses. Clinical management was altered in 10 patients because of the microbiological reports and in four patients with negative microbiology (chi2 = 8.19; P < 0.01). CONCLUSION: The standard of initial assessment was high in this specialist unit, and the presence of two or more 'markers of severity' did help predict outcome. A simple change in work practice resulted in a significant improvement in the thoroughness of microbiological investigations.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗

PubMed2001

System, environmental, and policy changes: using the social-ecological model as a framework for evaluating nutrition education and social marketing programs with low-income audiences.

A variety of nutrition education interventions and social marketing initiatives are being used by the Food Stamp Program to improve food resource management, food safety, dietary quality, and food security for low-income households. The Social-Ecological Model is proposed as a theory-based framework to characterize the nature and results of interventions conducted through large public/private partnerships with the Food Stamp Program. In particular, this article suggests indicators and measures that lend themselves to the pooling of data across counties and states, with special emphasis on systems, environment, and public policy change within organizations at the community and state levels.

Food Services↗

PubMed2004

Using medical record reminders to improve pneumococcal vaccination rates.

BACKGROUND: Despite widespread recommendations, the pneumococcal vaccination rate of patients 65 years of age or older is below target goals. A reminder system was instituted at the time of the scheduled primary care visit as a pilot quality improvement project. METHODS: Medical records staff used clinic schedules to identify patients 65 years of age or older, stamping that day's progress note with a reminder about pneumococcal vaccination. The physician could note previous vaccination or could order vaccination at the visit. If the patient was previously vaccinated or received a vaccine during that visit, an indicator was placed on the front of the chart. Staff at both clinics received an educational session on the indications and benefits of pneumococcal vaccination. RESULTS: Before implementing the reminder intervention, the pneumococcal immunization recording was comparable at the reminder clinic and at a second clinic that did not use reminders (x2 = 0.324, NS). At 6 months, 95/112 (85%) and 75/115 (65%) patients at the reminder and the second clinic had documentation of pneumococcal vaccination (x2 = 12.66, p < .005); at 12 months, 76% and 58% had such documentation. At the reminder clinic, the reminder stamp was found in 158/205 (77%) of the charts reviewed, with documentation of pneumococcal vaccination in 140/158 (89%) of these charts. DISCUSSION: The simple and inexpensive method of placing selective reminders in charts for elderly patients eligible for pneumococcal vaccination was highly effective in increasing the pneumococcal vaccination rate.

Florida↗

PubMed2003

Molding of three-dimensional microstructures of gels.

This Communication describes the use of patterned elastomeric stamps to mold, release, and stack hydrogels into three-dimensional microstructures. Molding of gels against stamps derivatized by a hexa(ethylene glycol)-terminated self-assembled monolayer or by an adsorbed monolayer of bovine serum albumin allowed the application of several soft lithographic techniques (replica molding, microtransfer molding, and micromolding in capillaries) to the microfabrication of gels. We describe procedures to generate coplanar or bilayered composites of gels.

Dimethylpolysiloxanes↗

PubMed2004

Edge spreading lithography and its application to the fabrication of mesoscopic gold and silver rings.

A new form of edge lithography, edge spreading lithography (ESL), has been demonstrated and applied to the formation of coinage metal rings. In this process, alkanethiols are delivered from a flat PDMS stamp to the surface of a metal film through a two-dimensional array of spherical silica colloids. The thiols further spread on the metal surface, forming highly ordered SAMs in the form of a ring pattern. Following lift-off of beads, the pattern in the SAMs can be transferred into the metal film through wet chemical etching, with SAMs serving as the resist. The dimensions of the rings can be readily controlled by several parameters such as the beads diameter, the concentration of the thiol solution, and the contact time between the stamp and the silica beads.

Journal Article↗

PubMed2005

Combining microfluidic networks and peptide arrays for multi-enzyme assays.

This paper reports the use of microfluidic networks (muFNs) to both prepare peptide microarrays and carry out label-free enzyme assays on self-assembled monolayers (SAMs) of alkanethiolates on gold. A poly(dimethylsiloxane) (PDMS) stamp fabricated with microchannels is used to immobilize a linear array of cysteine-terminated peptides onto SAMs presenting maleimide groups. The stamp is then reapplied to the SAM in a perpendicular direction to introduce enzyme solutions so that each solution can interact with an identical linear array of immobilized peptides. The muFNs enable multiple enzyme-substrate interactions to be simultaneously evaluated at a submicroliter scale, while the use of SAMs enables the use of MALDI mass spectrometry (MS) to analyze the enzyme activities. This paper demonstrates applications of this system for assaying multiple kinases and for profiling the activities of kinases and phosphatases in human K562 cell extracts. The combination of muFN, SAMs, and MS detection provides a flexible platform for assaying enzyme activities in biological samples.

Amino Acid Sequence↗

PubMed2006

High-efficiency stepwise contraction and adsorption nanolithography.

A new miniaturization protocol is demonstrated using stretching and relaxation of an elastomer substrate. A designed microstructure is formed on the stretched substrate and subsequently becomes miniaturized when the substrate relaxes. More importantly, the miniaturized structures can be transferred onto a new substrate for further miniaturization or can be utilized as stamps for nanolithography of designated materials. As an example of this approach, an elastic mold was first cast from a Si mold containing periodic line arrays of 1.5-microm line width. Upon relaxation, line width is reduced to 240 nm. The new elastomer may be used as stamps for micro- and nanofabrication of materials such as proteins. The polymer surface roughness or wrinkling behavior at nanoscale is found to follow classic stability model in solid mechanics. This observation provides means to design and control the surface roughness to meet specific requirements.

Acrylic Resins↗

PubMed2004

Patterned and controlled polyelectrolyte fractal growth and aggregations.

Two-dimensional patterned and controlled polyelectrolyte aggregations (e.g., tree-like ramified structures) created by microcontact printing have been demonstrated and discussed. Polyelectrolyte-micropatterned aggregations on surfaces were controlled by the micropattern size and shape of PDMS stamps. The formation of aggregates was dependent on the ink and surface conditions, and the aggregates consisted of two distinct layers; strongly adsorbed, primary uniform layers and weakly adsorbed, secondary aggregation layers positioned on top of the primary layers. The adsorption of the primary layers was strong enough not to be washed away, while the aggregated secondary layers were easily removed by washing. The aggregation of secondary layers showed typical tree-like ramified structures of fractal growth and aggregation. Directional and confined stamping led to directing and confining the growth of the fractal polyelectrolyte clusters, respectively. The micropatterned primary uniform layers were not removed by extensive washing, and they were identified by selective nickel plating and charged particle selective adsorption in which the surface formed positive and negative micropatterns. These functional and patterned surfaces have great potentials for advanced devices and sensors.

Journal Article↗

PubMed2004

Spreading of 16-mercaptohexadecanoic acid in microcontact printing.

Spreading in microcontact printing refers to the process or processes by which the ink molecules end up in the parts of the substrate that are adjacent to the contacted areas but which are not contacted themselves. This has been investigated for different inking concentrations of 16-mercaptohexadecanoic acid (MHDA). Spreading of MHDA takes place with retention of a well-defined demarcation. Feature sizes can be controlled by varying the contact times. Spreading, however, only takes place beyond a certain threshold concentration. For low ink concentrations the edges of stamp features dominate the ink transfer. For these low concentrations the extent of this edge dominance depends strongly on ink concentration rather than on contact time. These observations indicate a dominant role of the stamp surface in the processes of pattern formation and spreading.

Journal Article↗

PubMed2005

One-step multilevel microfabrication by reaction--diffusion.

A new experimental technique is described that uses reaction--diffusion phenomena as a means of one-step microfabrication of complex, multilevel surface reliefs. Thin films of dry gelatin doped with potassium hexacyanoferrate are chemically micropatterned with a solution of silver nitrate delivered from an agarose stamp. Precipitation reaction between the two salts causes the surface to deform. The mechanism of surface deformation is shown to involve a sequence of reactions, diffusion, and gel swelling/contraction. This mechanism is established experimentally and provides a basis of a theoretical lattice-gas model that allows prediction surface topographies emerging from arbitrary geometries of the stamped features. The usefulness of the technique is demonstrated by using it to rapidly prepare two types of mold for passive microfluidic mixers.

Journal Article↗

PubMed2004

Patterning adjacent supported lipid bilayers of desired composition to investigate receptor-ligand binding under shear flow.

To achieve efficient targeting, carriers containing either drugs or imaging agents must have surface properties that promote binding to targets yet at the same time block rapid immune system clearance. Here we describe a versatile technique that allows simultaneous comparison of the effects of carrier surface composition on binding properties under identical flow conditions. Parallel lanes of supported lipid bilayers that mimic the surface of liposomal delivery vehicles are formed using the vesicle fusion method in microfluidic channels created via standard soft lithography techniques. Vesicle stock solutions are premixed and injected into lanes formed by a poly(dimethylsiloxane) (PDMS) stamp reversibly sealed to a glass slide to create adjacent lanes of distinct composition. After removing the stamp, an adsorbed layer of bovine serum albumin (BSA) is used to prevent bilayer spreading before assembling the patterned substrate into a flow chamber for binding studies. Advantages of this method include easy and rapid preparation of bilayers with desired compositions from an unlimited number of lipid types, choice of feature size, time-stable features, and low nonspecific binding. Feature sizes on the order of tens of microns allow multiple compositions to be analyzed in one field of view, thereby reducing the number of experiments, ensuring identical flow conditions, and enabling simultaneous incorporation of controls. We show that the presence of a long poly(ethylene glycol) (PEG) tether (MW 2000) between the lipid and ligand results in higher detachment resistances as compared to a short six-carbon spacer.

Animals↗

PubMed2004

Structural characterization of microcontact printed arrays of hexa(ethylene glycol)-terminated alkanethiols on gold.

This paper reports on the structural characteristics of microcontact printed oligo(ethylene glycol)-terminated alkanethiol layers, HS(CH2)15CONH-(CH2CH2O)6-H (hereafter EG6), on gold. Microwetting, contact angle goniometry, imaging null ellipsometry, and infrared reflection-absorption spectroscopy (IRAS) are used to characterize the printed EG6 layers, and the quality of these layers in terms of layer thickness and the crystallinity of the alkyl and ethylene glycol portions is compared with data obtained from analogous layers prepared by solution self-assembly. The outcome of the printing process is critically dependent on the experimental parameters used to prepare the patterns. It is found that high quality layers, consisting of densely packed all-trans alkyl chains terminated with relatively helical hexa(ethylene glycol) tails, are formed by inking the poly(dimethylsiloxane) (PDMS) stamp with a 1 mM EG6 solution and contacting it with gold for 15 min. The homogeneity of printed layers is not as good as the homogeneity of those prepared from solution under similar conditions, most likely because of simultaneous transfer of low molecular weight residues from the PDMS stamp. These residues, however, can be easily removed upon ultrasonication in ethanol without affecting the quality of the printed layer. Further on, the microscopic square-shaped bare gold patterns formed after microcontact printing (muCP) are subsequently filled with 16-hexadecanoic acid (hereafter THA) or HS(CH2)15CONH-(CH2CH2O)6-COOH (hereafter EG6COOH) to provide a microarray platform for further covalent attachment of biomolecules. Well-defined structures in terms of wettability contrast, sharpness, and height differences between the printed and back-filled areas are confirmed by imaging null ellipsometry and microscopic wetting.

Gold↗

PubMed2006

Scanning force microscopy and fluorescence microscopy of microcontact printed antibodies and antibody fragments.

Unlabeled primary immunoglobulin G (IgG) antibodies and its F(ab')2 and Fc fragments were attached to oxygen-plasma-cleaned glass substrates using either microcontact printing (MCP) or physical adsorption during bath application from dilute solutions. Fluorescently labeled secondary IgGs were then bound to surface-immobilized IgG, and the relative surface coverage was determined by measuring the fluorescence intensity. Results indicated that the surface coverage of IgG increased with increasing protein solution concentration for both MCP and bath-applied IgG and that a greater concentration of IgG was transferred to a glass substrate using MCP than during physisorption during bath applications. Scanning force microscopy (SFM) showed that patterned MCP IgG monolayers were 5 nm in height, indicating that IgG molecules lie flat on the substrate. After incubation with a secondary IgG, the overall line thickness increased to around 15 nm, indicating that the secondary IgG was in a more vertical orientation with respect to the substrate. The surface roughness of these MCP patterned IgG bilayers as measured by SFM was observed to increase with increasing surface coverage. Physisorption of IgG to both unmodified patterned polydimethylsiloxane (PDMS) stamps and plasma-cleaned glass substrates was modeled by Langmuir adsorption kinetics yielding IgG binding constants of K(MCP) = 1.7(2) x 10(7) M(-1) and K(bath) = 7.8(7) x 10(5) M(-1), respectively. MCP experiments involving primary F(ab')2 and Fc fragments incubated in fluorescently labeled fragment-specific secondary IgGs were carried out to test for the function and orientation of IgG. Finally, possible origins of MCP stamping defects such as pits, pull outs, droplets, and reverse protein transfer are discussed.

Adsorption↗

PubMed2006

Surface energy modified chips for detection of conformational states and enzymatic activity in biomolecules.

A novel patterning method for anchoring biomolecules and noncovalent assembled conjugated polyelectrolyte (CPE)/biomolecule complexes to a chip surface is presented. The surface energy of a hydrophilic substrate is modified using an elastomeric poly(dimethylsiloxane) (PDMS) stamp, containing a relief pattern. Modification takes place on the parts where the PDMS stamp is in conformal contact with the substrate and leaves low molecular weight PDMS residues on the surface resulting in a hydrophobic modification, and then biomolecules and CPE/biomolecule complexes are then adsorbed in a specific pattern. The method constitutes a discrimination system for different conformations in biomolecules using CPEs as reporters and the PDMS modified substrates as the discriminator. Detection of different conformations in two biomacromolecules, a synthetic peptide (JR2E) and a protein (calmodulin), reported by the CPE and resolved by fluorescence was demonstrated. Also, excellent enzyme activity in patterned CPE/horseradish peroxidase (HRP) enzyme was shown, demonstrating that this method can be used to pattern biomolecules with their activity retained. The method presented could be useful in various biochip applications, such as analyzing proteins and peptides in large-scale production, in making metabolic chips, and for making multi-microarrays.

Calmodulin↗

PubMed2005

Supramolecular nanostamping: using DNA as movable type.

Here we present a novel printing technique (that we call supramolecular nanostamping), based on the replication of single-stranded DNA features through a hybridization-contact-dehybridization cycle. On a surface containing features each made of single-stranded DNA molecules of known sequence, the complementary DNA molecules are hybridized, spontaneously assembling onto the original pattern due to sequence-specific interactions. These complementary DNA strands, on the end that is assembled far from the original surface, are 5' modified with chemical groups ("sticky ends") that can form bonds with a target surface that is brought into contact. Heating induces dehybridization between DNA strands, leaving the original pattern on the original surface and the copied pattern on the secondary substrate, and thus stamping (see Figure 1). Molecular recognition provides the unique and disruptive ability of transferring large amounts of information in a single printing cycle, that is the simultaneous stamping of spatial information (i.e., the patterns) and of chemical information (i.e., the features' DNA sequence--their chemical composition). This method combines high resolution (<40 nm) with the advantage of an exponential increase in the number of masters; in fact, any printed substrate can be reused as a master. Patterns fabricated via very different lithographic techniques can be replicated.

DNA↗