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PubMed1999

Separation of double-stranded DNA fragments in plastic capillary electrophoresis chips by using E99P69E99 as separation medium.

The separation of double-stranded DNA (dsDNA) fragments in polymethylmethacrylate (PMMA) capillary electrophoresis (CE) chips by using E99P69E99 as a separation medium has been demonstrated. The PMMA CE chips were simply manufactured by micromachining and adhesive tape sealing. To make the separation channel compatible with the separation medium, a dynamic nonionic surfactant coating procedure was developed, which made the plastic separation channel sufficiently hydrophilic to allow the separation medium to fill the channel by capillary action. Subsequent separation of DNA fragments was successful with a separation efficiency of the order of 10(4) theoretical plates over an effective separation distance of 1.5 cm. By using an applied electric field strength of 200 V/cm, the separation of low DNA mass ladder was completed within 5 min. The simple coating procedure, together with the self-assembled viscosity-adjustable separation medium, should be useful to meet some of the essential requirements for developing single-use disposable CE chips. Coating the channels with polymer blends of PMMA and the separation medium also showed promise.

Animals↗
PubMed1977

A new simple method for canalicular intubation.

A series of ten patients had canalicular intubation with Teflon and Silastic tubing. Our experience has shown that Silastic tubing appears to be more pliable, has good capillary action, and causes less irritation within the canalicular system than polyethylene or Teflon tubing. Five patients had very good results, three patients were improved, and there were two failures. We believe that this technique is useful in selected cases where there is an obstructed canaliculus near its junction with the sac. There should be only minimal scarring in the medial canthal area for the greatest chance of success with this method.

Dacryocystorhinostomy↗
PubMed1999

Immunochromatographic membrane strip assay system for a single-class plasma lipoprotein cholesterol, exemplified by high-density lipoprotein cholesterol measurement.

In assessing risk factors of coronary heart disease, a membrane immunochromatographic system that minimizes requirements of instrument and reagent handling was investigated by utilizing high-density lipoprotein (HDL) cholesterol (HDL-C) as model analyte. The system is composed of four functional membrane strip pads connected in sequence as follows (from the bottom): immunoseparation based on the biotin-streptavidin reaction; catalytic conversion of cholesterol to hydrogen peroxide; production of a colorimetric signal; and induction of a continuous wicking of medium. For immunochromatography, a monoclonal antibody, specific to apolipoprotein B100 that is present on the surfaces of low-density lipoproteins (LDL) and very low-density lipoproteins (VLDL), with a high binding constant (5 x 10(10) L/mol), was raised and chemically conjugated to streptavidin. The conjugate was first reacted with lipoprotein particles, and this mixture was absorbed by the capillary action into the biotin pad of the system. After being transferred by medium, immunocapture of LDL and VLDL particles onto the biotin pad took place, and in situ generation of a colorimetric signal in proportion to HDL-C occurred consecutively. The capture was selective as well as effective (minimum 88% of LDL and VLDL in clinical concentration ranges), and the detection limit of the HDL-C was far lower than 20 mg per 100 mL. The same concept may also be applicable to LDL cholesterol measurement provided suitable antibodies specific to HDL and VLDL are available.

Animals↗
PubMed1999

Failure of a new double glove hole detection system in the Emergency Department.

The purpose of this study was to evaluate the performance of a new double glove hole detection system in the Emergency Department. First, the frequency of holes in both gloves of the double glove hole detection system was determined using a watertight test method. Second, the frequency of glove puncture was determined first by searching for the optical color change that occurs with the ingress of fluid in the double glove hole detection system. These same gloves were then removed and also checked for holes by the watertight test method. After removal from the package, no holes were detected in the two gloves of the system using the watertight test method. In 50 consecutive patients, there was no color change in the inner glove indicating glove puncture. When these same gloves were then tested with the watertight test method, 14 of the 50 double glove hole detection systems failed; all 14 outer gloves were punctured, and three of the inner gloves had holes without demonstrable injury to the skin. This double glove hole detection system is not a reliable system to detect holes in relatively dry clinical settings because the ingress of fluid by capillary action between the gloves is necessary to cause a color change in the inner glove that signals the presence of a hole.

Arthroplasty↗
PubMed1994

A rapid simple in situ hybridization method for herpes simplex virus employing a synthetic biotin-labeled oligonucleotide probe: a comparison with immunohistochemical methods for HSV detection.

We examined 28 paraffin-embedded tissue specimens with histologic evidence of herpes virus infection by in situ hybridization (ISH) utilizing manual capillary action technology (MicroProbe Staining System) and a 21 base synthetic multibiotinylated oligonucleotide probe from the HSV glycoprotein C region. The results were compared to a rapid simple immunohistochemical (IHC) protocol for detection of HSV proteins. HSV was detected by ISH and IHC in all but one specimen which was shown to be positive for varicella zoster virus by direct fluorescent antibody studies. Hybridization signal was confined to the nucleus in all cases. Staining was identified in cells with early as well as late cytopathic effect. IHC produced intense nuclear and/or cytoplasmic signal in infected cells and stained in areas of necrosis which were otherwise spared by ISH. HSV was detected by IHC and/or ISH in 3/5 specimens with histology suggestive of, but not diagnostic for, HSV infection. Both techniques were sensitive and specific for HSV, resulted in rapid detection of the pathogen in routinely processed tissues, and may be useful in cases where the histologic impression is equivocal for HSV infection. ISH for HSV may be preferred because it can identify early HSV infection, which in turn can be treated with antiviral agents.

Base Sequence↗
PubMed1999

Liposome dehydration on nitrocellulose and its application in a biotin immunoassay.

The feasibility of utilizing dehydrated liposomes in the development of a simple immunoassay device for point-of-care diagnostics or field assays was demonstrated. The recovery of liposomes after a cycle of dehydration and rehydration was studied using biotin-tagged, dye-loaded liposomes with antibiotin antibodies immobilized in a defined zone on nitrocellulose strips. Liposomes were vacuum-dehydrated on the strip at a location below the antibiotin zone. The strip was placed in a tube containing a carrier solution and capillary action brought the solution to the dehydrated liposomes, rehydrated them, and caused them to migrate to the antibody zone where intact liposomes were captured and measured optically. High concentrations of either trehalose or sucrose external to the liposomes and both polyvinylpyrrolidone and gelatin in the membrane blocking reagent were essential for preservation of the dehydrated/rehydrated liposomes on nitrocellulose. Between 70 and 80% of the liposomes were recovered on the nitrocellulose strips after a cycle of dehydration and rehydration. The dehydrated liposomes on the strips were stable for at least 1 year when stored in vacuum-sealed plastic bags at 4 degrees C. The technique was successfully applied to the development of a rapid one-step strip immunoassay for biotin.

Biotin↗
PubMed1999

Pendular, Funicular, and Capillary Bridges: Results for Two Dimensions.

Theconfigurations and interactions in a system of particles connected by liquid bridges are investigated. The basic phenomena have been elucidated using two-dimensional models with equal sized particles. With this restriction all possible contact configurations can be obtained by considering three particles. Different configurations of the liquid distribution can occur depending on the amount of liquid. For small quantities of interstitial liquid, the only possible configurations are pendular bridges. For large amounts of liquid a fully saturated state called the capillary state can exist. There are also a number of intermediate cases characterized by varying degrees of interstitial voidage, which are classified as funicular configurations. The interactions of the particles by means of the interstitial liquid binder in various possible configurations have been characterized, and the interparticle forces and the energy of the system due to the capillary action of the liquid bridges have been calculated. The changes in configuration, force, and energy as one particle is pulled away are also considered. Since the parametric space for calculation is large, only two representative cases are presented. Copyright 1999 Academic Press.

Journal Article↗
PubMed2000

Development of rapid one-step immunochromatographic assay.

An analytical system for a one-step immunoassay has been constructed using the concept of immunochromatography. The system employed two different antibodies that bound distinct epitopes of an analyte molecule: an antibody labeled with a signal generator (e.g., colloidal gold), which was placed in the dry state at a predetermined site on a glass-fiber membrane, and another antibody immobilized on the surface of a nitrocellulose membrane. Three membranes, one with the tracer, one with immobilized antibody, and a cellulose membrane as the absorbent of medium (in a sequence from the bottom), were attached to a plastic film and cut into strips. Aqueous medium containing analyte absorbed from the bottom end of the immunostrip dissolved the labeled antibody, and the antigen-antibody binding complex formed was transported into the next nitrocellulose membrane by the flow caused by capillary action. The complex subsequently reacted with the immobilized antibody, which generated a signal in proportion to the analyte concentration. The convective mass transfer of the immunoreactant to the binding partner allowed the assay to be performed with no handling of reagents. The reaction, however, was carried out under nonequilibrium conditions, which resulted in decreased sensitivity as compared with assays performed in an equilibrium mode (e.g., ELISA). To minimize such sacrifice, major factors that control system performance were identified and the system was then devised under optimal conditions.

Animals↗
PubMed1986

Investigation of the interaction of cardiotoxic anticancer agents using the fetal mouse heart organ culture system.

The fetal mouse heart organ culture system was utilized in an effort to document and predict the potential cardiotoxic effects of ionizing radiation, Adriamycin (ADR), and Dihydroxyanthraquinone (DHAQ); alone and in combination. These antineoplastic agents have been shown to produce clinical cardiomyopathy which is often dose-limiting. Fetal mouse hearts (gestational day 17) were removed and placed in a culture system of 6-well microtiter plates. A single heart was placed in each well on a piece of aluminium mesh, above the culture medium but bathed by capillary action. The plates were then placed in a 100% oxygen environment and incubated at 37 degrees C. Treatments performed on day 1 after culture were Cs-137 irradiation (10, 20, or 40 Gy); ADR (10, 30, or 100 micrograms/ml); DHAQ (5, 20, or 50 micrograms/ml); or various combinations of drugs and radiation. Hearts were checked every day for functional activity as evidenced by continuous heart best. Untreated hearts beat rhythmically for up to 9 days (average = 6.8 days); treated hearts stopped beating between 2 and 7 days after treatment. Using this endpoint of functional retention time (FRT), dose response curves were obtained for all individual agents. Combinations of ADR and DHAQ (at concentrations that resulted in FRTs of 3.5 days) produced no greater effect than either agent alone. However, the combination of radiation (FRT = 5.3 days) with ADR, DHAQ or both drugs was more effective than was drug alone. This system may help to predict the cardiotoxic effects that result from the use of these drugs and radiation.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMedDate not supplied

Non-penetrating deep sclerectomy (NPDS) with or without collagen device (CD) in primary open-angle glaucoma: middle-term retrospective study.

The purpose of this study is to evaluate the middle-term tonometric results of a new filtering procedure, the nonpenetrating deep sclerectomy with or without collagen device, in primary open-angle glaucoma. This technic aims to eliminate or minimize the complications of classical trabeculectomy. METHOD: This procedure was carried out by Koslov and colleagues. This is performed under a limbal-based conjunctival flap and a superfical scleral flap, the ablation of a deep scleral flap taking away the external wall of Schlemm's canal, only living in place the Descemet membrane. One must obtain a visible filtration across the opened Schlemm's canal and Descemet membrane. To improve the aqueous filtration, a cylindric collagen device, made from porcine scleral tissue, biocompatible, known for its high water content, is fixed in the deep scleral bed with a 10/0 nylon suture. This device provides a support for the elimination route of aqueous humor and acts like a sponge, carrying the liquid by capillary action. It is sterilized by irradiation. Full guarantee against viral contamination is provided. This procedure ends in one suture (10/0 nylon) of superficial scleral flap and conjunctival closing suture. When NPDS is performed without CD a sponge of 5FU is used and the superficial scleral flap is not sutured. RETROSPECTIVE STUDY: Our material included 111 patients, 148 eyes in CD group; 43 patients, 55 eyes in the group without CD. The average follow-up was 13.3 +/- 5.8 months in the CD group, 7.2 +/- 3.5 months in the group without CD. All patients presented a POAG without risk factors of bleb failure. RESULTS: The delta average IOP before the operation and at the end of the follow-up period was 7.2 +/- 6.3 mmHg in the CD group; 8.3 +/- 7.6 mmHg in the group without CD (no significant difference). The probability-success rate with the Kaplan-Meier method (IOP < or = 20 mmHg) was, in the CD group, at 18 months, 68% and 69% in the group without CD, without medical treatment. With monotherapy, the success rate was 85% in the CD group, 74% in the group without CD (p < or = 0.05). PROSPECTIVE STUDY: Afterwards, we have conducted a prospective study comparing two groups of patients with POAG without risk factors of bleb failure, operated with and without collagen device, without 5FU in the second group. Our material included 31 patients, 31 eyes, one eye for each patient, two surgeons; 17 eyes in the CD group, 14 eyes in the group without CD. The average age was 65.8 +/- 8.2 years in the first group; 64.1 +/- 10.3 in the second group. The average follow-up was 11 months in both groups. RESULTS: delta average IOP was 8.3 +/- 5.8 in the CD group; 12.3 +/- 6 in the group without CD (p < 0.05). The probability-success rate without treatment at 12 months: 58% in the first group, 90% in the second group (p < 0.05) and with monotherapy: 80% and 90% (N.S.). In both studies, in both groups, except microperforations, more frequent in the prospective group without CD, no complications of the trabeculectomy were observed. The mean change in visual acuity was inferior to 0.1 at the end of the follow-up. A postoperative rise in IOP can occur. It can be due to an internal obstruction (goniosynechiae or bad filtration). It can be treated with Nd-Yag laser. It can also be due to external obstruction, treated by 5FU injections into the bleb. The success of these procedures were similar in the whole group. CONCLUSION: Non penetrating deep sclerectomy can be considered as an excellent alternative to trabeculectomy in open and wide angles. It does not modify visual acuity. It carries away less complications than trabeculectomy and the use of antimitotic agents is safer. Collagen device does not seem, at middle-term, to improve tonometric results.

Adult↗
PubMed1989

Evaluation of a rapid latex agglutination test for detection of group B streptococci in vaginal specimens.

A rapid latex agglutination test (Bactigen Group B Streptococcus Cervical Screen) for detection of group B streptococci in cervical-vaginal specimens was evaluated using two different slide systems, the traditional serologic slide and capillary action track (Trak) slide. Culture was used as reference method. A total of 344 cervical-vaginal specimens were tested. The group B streptococci carrier rate was found by culture to be 10.8%, 56.8% of these specimens being heavily colonized. The sensitivity and specificity of the latex agglutination test in heavily colonized specimens was 95.2% and 99.3% for the serologic and track slides respectively. The overall sensitivity, including lightly colonized specimens, was 62.2%. The positive predictive value was 92% for both slide systems, and the negative predictive value 95.4% and 95.6% for the serologic and track slides respectively. The latex agglutination test, used with either slide, provides a rapid and effective method for identification of specimens heavily colonized with group B streptococci. The track slide may provide a convenient alternative to serologic slides since it does not require rotation.

Cervix Uteri↗
PubMed2001

Chip electrochromatography of polycyclic aromatic hydrocarbons on an acrylate-based UV-initiated porous polymer monolith.

The first rigorous evaluation of a UV-initiated porous polymer monolith (PPM) as a stationary phase for chip electrochromatography (ChEC) is described. All channels in an offset T-injector-design-chip (25-microm deep by 50-microm wide channels) were filled by capillary action with an acrylate-based PPM precursor solution and polymerized in situ using 365 nm light for several minutes. Photodefinability of the monolith cast in the channels during the polymerization process was also demonstrated by masking off the injection arms during photoinitiation. The chromatographic performance of this chip was compared with that of chips completely filled with monolith. The detection window was photodefined after polymerization using the detection laser (257 nm doubled argon ion laser) to depolymerize the detection window. A successful ChEC separation of 10 out of 13 polycyclic aromatic hydrocarbons (PAH) was performed with on-column, off-packing laser-induced fluorescence detection at 257 nm. Van Deemter plots for early-, middle-, and late-eluting compounds showed the minimum plate height to be 5 microm. The average number of theoretical plates per meter for the PAH was 200,000. Several factors contributed to irreproducible results. Oxygen was observed to dynamically quench the fluorescence of the sample over time. Improved sealing of the reservoirs solved this problem. A within-chip variability in the retention time of 2-10% RSD was observed. These results demonstrate the feasibility and reliability of the PPM as a solid reversed-phase for electroosmotic flow-driven chip-based chromatography in microscale total analysis systems.

Journal Article↗
PubMed2003

Scanning electron microscopic analysis of the mineralization of type I collagen via a polymer-induced liquid-precursor (PILP) process.

We have put forth the hypothesis that collagen is mineralized during bone formation by means of a polymer-induced liquid-precursor (PILP) process, in which a liquid-phase mineral precursor could be drawn into the gaps and grooves of the collagen fibrils by capillary action, and upon solidification, leave the collagenous matrix embedded with nanoscopic crystallites of hydroxyapatite. This hypothesis is based upon our observations of capillarity seen for liquid-phase mineral precursors generated with calcium carbonate. Here, we demonstrate proof-of-concept of this mechanism by mineralizing Cellagen sponges (type I reconstituted bovine collagen) in the presence of a liquid-precursor phase to calcium carbonate. Scanning electron microscopy (SEM) was used to examine the mineralized collagen, which in combination with selective etching studies, revealed the extent to which the mineral phase infiltrated the collagenous matrix. A roughly periodic array of disk-like crystals was found to be embedded within the collagen fibers, demonstrating that the mineral phase spans across the diameter of the fibers. Some of the morphological features of the mineralized fibers in our in vitro model system are similar to those seen in natural bone (albeit of a different mineral phase), lending support to our hypothesis that these non-equilibrium morphologies might be generated by a PILP process. SEM provides a different perspective on the morphology of bone, and has been useful here for examining the extent of mineralization in composite structures generated via the PILP process. However, further investigation is needed to examine the nanostructural arrangement of the crystallites embedded within the collagenous matrix.

Animals↗
PubMed1999

An improved method for STR analysis of bloodstained denim.

Indigo dye is used to dye denim and other fabrics. It is now accepted that if this is co-extracted with the DNA, it may inhibit PCR amplification. A simple, improved method is described for the extraction of DNA from bloodstained denim for PCR amplification and short tandem repeat (STR) analysis. The DNA was extracted by constructing a blotting system using capillary action to draw a saline solution through the denim. The transferred material was collected onto nylon membranes and these were processed by chelex extraction. A variety of coloured denim substrates and other heavily dyed fabrics, including case work samples were used. In all cases the DNA was extracted, amplified and typed correctly.

Alleles↗
PubMed1991

Development of an automatic machine for in situ hybridization and immunohistochemistry.

An instrument for the automation of in situ hybridization and immunohistochemistry has been developed. This machine is capable of analyzing 20 microscope glass slides via all of the steps required for colorimetric in situ hybridization or immunohistochemistry. The slides are placed specimen-side down on a specialized Teflon slide-holder set in the reaction chamber of the machine. The system uses a unique type of capillary action between the slide and the holder. The holder has two small holes and is designed to apply, incubate and sequentially add and remove reagents from the slide surface. The system performs the complete processes of in situ hybridization and immunohistochemistry from dewaxing to colorization. Some applications were carried out using this instrument. Cultured cells infected with cytomegalovirus, adenovirus, or herpes simplex virus were hybridized with homologous biotinylated probes, and showed strong purple signals with alkaline phosphatase in the presence of nitroblue tetrazolium and 5-bromo-4-chloro-3-indolyl phosphate. Automatic in situ hybridization using other colorimetric detection systems (e.g., peroxidase-labeled probes/diaminobenzidine/H2O2) was also examined in cells infected with Chlamydia trachomatis and in paraffin-embedded hepatic tissue sections from patients with hepatitis. For conventional immunohistochemical staining, formalin-fixed and paraffin-embedded tissues were used. Glial fibrillary acidic protein and gamma-immunoglobulins were detected automatically in human brain white matter and tonsillar tissues, respectively, as peroxidase-based reddish signals. The intensity of staining was equal to that achieved by manual methods.

Automation↗
PubMedDate not supplied

Planar chromatography: current status and future perspectives in pharmaceutical analysis--I. Applicability, quantitation and validation.

The most important features of capillary action planar chromatographic methods are summarized. Certain properties such as ease in operation, sensitivity to experimental conditions, stationary and mobile phase selection, phase system optimization, separation efficiency and detection possibilities are discussed only briefly. Other aspects such as the applicability of planar chromatographic methods in pharmaceutical analysis, quantitation and validation are considered in more detail. The advantages and limitations of different evaluation techniques (visual comparison, spot elution techniques and in situ densitometry) are also discussed. Validation steps specific to planar chromatographic methods are emphasized.

Chemistry, Pharmaceutical↗
PubMed2006

Dry reagent dipstick test combined with 23S rRNA PCR for molecular diagnosis of bacterial infection in arthroplasty.

Periprosthetic joint infections present a challenging problem in orthopaedics. Conventional methods for detection of arthroplasty infections rely on bacterial culture of synovial fluid aspirates. During recent years, however, molecular tests that are based on DNA amplification by the polymerase chain reaction (PCR), followed by electrophoretic analysis of the products, have been introduced. We report a simple and inexpensive assay that allows visual detection and confirmation of the PCR-amplified sequences by hybridization within minutes. The assay is performed in a dry reagent dipstick format (strip) and does not require special instrumentation. Universal primers are used for PCR of the 23S ribosomal RNA (rRNA) gene. The biotinylated amplification product is hybridized with dA-tailed probes that are specific for six pathogens commonly involved in periprosthetic joint infections. The mixture is applied to the strip, which is then immersed in the appropriate buffer. The buffer migrates along the strip by capillary action and rehydrates gold nanoparticles with oligo(dT) strands attached to their surface. The nanoparticles bind to the target DNA through hybridization, and the hybrids are captured by immobilized streptavidin at the test zone of the strip, producing a characteristic red line. Unbound nanoparticles are captured by immobilized oligo(dT) strands at the control zone of the strip, generating a second line. The dipstick test was applied to the detection of Escherichia coli, Staphylococcus aureus, Staphylococcus epidermidis, Streptococcus pneumoniae, Enterococcus faesium, and Haemophilus influenza. Twelve samples of synovial fluids from patients were analyzed for the detection and identification of the infection caused by the six pathogens. The results were compared with bacterial cultures.

Arthroplasty↗
PubMed2004

Organic plasma process for simple and substrate-independent surface modification of polymeric BioMEMS devices.

A polymeric bio micro electromechanical systems (BioMEMS) device was fabricated using organic plasma polymerization, by which the surface of a polymeric substrate could easily be modified through vapor-phase deposition of organic thin films. This technique, capable of polymeric deposition of any kind of monomer, can serve the purpose of anti-fouling coating, wettability control, or layer-to-layer interface creation, on the surface of any given chemically-inert polymeric substrate without involving cumbersome surface organic reactions. A prototype device was fabricated to have an array of electrochemical glucose biosensors with the three electrode configuration, each of which has a microfluidic channel (500 microm x 800 microm) for capillary-action-driven sample delivery and the concerned enzymatic reaction. Stressing the advantages of the plasma polymerization process using a polymeric substrate together with some additional features accomplished in our device fabrication, new possibilities in the field of polymeric BioMEMS are discussed.

Biosensing Techniques↗