A method for administration of prolonged intravenous infusion of prostacyclin (PGI2) to unanesthetized rats.

Prostacyclin is short acting and chemically unstable. To study sustained effects in an intact animal, prolonged intravenous infusion may be required. The compound has adequate stability for 24 hr in pH 10.0 carbonate buffer at 0 degrees. A "displacement syringe" is described wherein the prostacyclin solution is stored in a rubber bag inside the barrel of a 5 ml syringe. This device is placed in an ice bath. A syringe pump drives water into the barrel displacing an equal volume of solution out of the bag. Chronic venous cannules, saddles, and flow-through swivels are used as for drug self-administration studies. A simple, inexpensive rack for use with conventional individual hanging cages is also described.     

Development of Biotechnology Products in Pre-filled Syringes: Technical Considerations and Approaches

A monoclonal antibody (mAb) product development case study is presented to address some of the issues faced during developing a pre-filled syringe (PFS) product for a biotherapeutic. In particular, issues involving incompatibility with silicone oil and a stability-based approach for selection of PFS barrel and tip cap components have been discussed. Silicone spiking studies followed by exposure to agitation stress or accelerated temperature conditions were used to check for incompatibilities of the mAb with silicone oil, a necessary product contact material in PFS. In addition, screening studies to compare various closure materials as well as syringe barrel processing methods were used to select the optimum closure materials as well as the correct syringe processing method. Results indicate that the model mAb formulation used was sensitive to high levels of silicone oil especially under accelerated temperature conditions resulting in formation of protein–silicone particles in the solution for samples that were spiked with the silicone oil. Agitation stress did not have any significant impact on the quality attributes tested. Samples stored in syringe barrels that were processed with sprayed-on silicone had higher levels of subvisible particles as compared to those that were processed with the baked-on process. The tip cap comparability study resulted in one tip cap material having superior compatibility among the three that were tested. The quality attribute that was most impacted by the tip cap materials was mAb oxidation. An approach for evaluation of primary packaging components during the development of pre-filled syringe presentations for biotechnology-based compounds has been highlighted.     

An Ideal Viscous Flow Porometer

A new design of viscous flow porometer is described which isbased on the pronciple of air suction into a syringe which thenmeasures the volume of air atmosphere pressure. This porometerallows accurate readings to be made on the 0-100 scale on thesyringe barrel when the piston has returned following its release.Each reading takes only seconds to make so that data can becollected for statistical analysis before stomata change inaperture and with minimal interference with the leaf envirnionment.Calibiration procedures are described: examples ofthe new porometersand also silicone rubber imprints support the validity of measurementsmade by the new porometer.     

树[鼠句]的基本实验操作方法

目的对树[鼠句]抓取和保定、采血、灌胃基本实验技术方法进行探讨,逐步规范树[鼠句]实验技术。方法选用成年树[鼠句]进行抓取和保定,分徒于法和器具法（自制捕捉保定袋）,对130只树[鼠句]采用尾静脉、股动（静）脉两种采血方法;采取人用8号胃管可对树[鼠句]绎口灌胃给药。结果所采用徒手、器具的方法抓取和保定树[鼠句]均能有效地控制动物,不会发生动物死亡或很少逃逸;两种方法都顺利采集到所需血量,股动（静）脉单次最大采血量可达2mL而不损伤动物;12只树[鼠句]连续灌胃10d,成功率100％。结论自制的捕捉保定袋经济实用,摸索的几种树[鼠句]实验技术方法具有操作简便、安全、快捷等优点。     

A novel concentric double-barrelled calcium-selective microelectrode for small cells

A novel concentric design of double-barrelled Ca2+-selective microelectrode, with an inner pipette tip that protrudes beyond an outer one, has recently been developed and is described. This configuration of pipettes was produced from concentric capillaries in one step using a horizontal pipette puller. For the tip of the inner barrel to protrude, Corning 1724 aluminosilicate glass was selected, as it has a higher melting point than the 1723 glass which is used for the outer barrel. To reduce electrode resistance the inner capillary was best made with a triangular shape. It was preferentially silanized in a dry box by injection of methyltrichlorosilane into only the inner barrel. The Ca2+ neutral carrier-based liquid membrane (ETH 1001) was back-filled from the tip to the shank of the inner pipette and above this CaCl2 solution was added. KCl, which contained EGTA and was buffered to pCa 7, was used to fill the reference barrel. These Ca2+ electrodes showed linear response with slope approximately equal to 30 mV for changes in Ca2+ concentration between 10(-3) and 10(-7) M in the presence of constant [K+]. They offer a number of advantages including a low noise level achieved by the presence of the external concentric KCl electrode, and a simple mechanical structure that allows applications to a variety of small cells.     

A modified blow-gun syringe for remote injection of captive wildlife

A modified syringe capable of automatic injection and suitable for use with a blow-gun is described. The syringe has been used successfully with white-tailed deer (Odocoileus virginianus) under confined conditions. Desirable characteristics for blow-gun syringes are discussed.     

Exponential gradient maker using a disposable syringe

With a simple modification, any disposable syringe can become a reliable and easy to use exponential gradient maker. The modification consists of two notches, made with a razor blade, in the borders of the rubber sealing tip of the plunger. A clamp in the tube connected to the syringe allows control over solution flow. With the clamp prohibiting drainage, the body of the syringe is filled with the desired volume of starting solution I. A magnetic stir bar, small enough to spin inside the syringe is included. The notched plunger is introduced until no air space remains. This forms the fixed volume, closed mixing chamber, while the rest of the volume of the syringe forms the open chamber. The two chambers are connected through the notches in the plunger. The ending solution II is poured after the introduction of the plunger. Opening the clamp allows solution I in the closed chamber to flow out, and the solution II in the open chamber flows through the notches and mixes with solution I. This exponential gradient maker can be reused many times, but the low cost of the components makes it potentially disposable. This feature is especially useful when using toxic chemicals, or when pouring polyacrylamide gradient gels, since the apparatus may be disposed of after contamination or eventual polymerization.     

A rapid, hygienic method for the preparation of fecal samples for liquid scintillation counting.

A method for the preparation of fecal samples for liquid scintillation counting is described which is rapid, hygienic, and inexpensive. By the use of a novel type of homogenizer, fecal samples can be homogenized while totally enclosed within a sealed, plastic bag, so reducing the possible risk of infection. The subsequent preparation of a clear solution suitable for liquid seintillation counting is performed using an “in-vial” digestion technique which enables any ¹⁴CO₂ released during digestion to be trapped within the vial.     

A simple automatic dispensing apparatus

A simple dispenser using a 2-ml disposable syringe is described. The range of operation is 0.02–1.5 ml and an unusual accuracy is obtained by employing a new principle for dispensers: the dispensed volume is only a fraction of total tidal flow of fluid in the dispenser.     

A simple device for the rapid concentration or isolation ofTetrahymena

We have developed a practical and economical device that efficiently concentrates ciliated cells, such asTetrahymena pyriformis, by using only low-speed, non-damaging centrifugation. This is carried out with a simple device consisting of a microcentrifuge tube connected to the barrel of a 60-ml plastic syringe. This device allows the concentration of essentially all the cells into a small volume without damaging cell structures or reducing their viability.     

A modified equilibrium dialysis method for studying fatty acid binding to proteins

A modified equilibrium dialysis method is described which is suitable for investigating the binding of fatty acids in the form of aqueous micellar dispersions to proteins. The method uses a permeant chromophore which complexes reversibly with free fatty acid within the dialysis bag. The concentration outside the dialysis bag is determined spectrophotometrically. Binding of oleic acid to bovine serum albumin is given as an example. A simplified analysis of fatty acid binding is given and used to indicate the potential of the method.     

Stabilization of alginate beads using radiation polymerized polyacrylamide.

A technique has been described for the stabilization of calcium alginate beads using radiation polymerized acrylamide. The technique involved dropping a mixture containing the cells (20%), sodium alginate (2%), acrylamide (2.5%) and N-N'-methylene-bis-acrylamide (0.1%) through a syringe needle into cold (-75 degrees C) toluene. The frozen beads obtained were exposed to 60Co gamma-rays (0.5 KGy) and were then thawed in 0.1 M CaCl2 solution. Unlike the calcium alginate beads the conjugate beads were not found to be dissolved when incubated in 3% trisodium citrate solution. Stabilized beads containing entrapped yeast cells could be reused for over 15 batches for the inversion of sucrose without loss in activity or chemical integrity of the beads.     

An improved method for assessing soil-fungus population density

Summary A rapid method for determining soil fungal populations is described. A 25-g equivalent of oven-dry soil, in 225 ml of water in a 500-ml aspirator bottle, is agitated on a magnetic stirrer and samples withdrawn with a syringe while the suspension is in motion. Individual drops are placed in petri dishes and mixed with a suitable culture medium. The average weight of soil per drop is determined and colony counts related to dry weight of soil as usual. This syringe method is simple, eliminates much of the error common in serial dilution and pipetting, and allows for a high degree of reproducibility and possible standardization.     

Preparation of leukocyte-poor platelet concentrates from buffy coats by a new type of quadruple bag

A new quadruple blood bag system for collection of platelet concentrates is described. The principle modification consists of a smaller (150 ml) buffy coat bag in conjunction with SAG-mannitol for red cell preservation. Platelet yield and function were comparable to conventional techniques, leukocyte contamination was lower (1.7 +/- 1.3 x 10(7) per unit). The system is recommended for specialized institutions with a high demand of blood components.     

Peptide mapping and microsequencing of proteins separated by SDS-PAGE after limited in situ acid hydrolysis.

A method is described for the isolation of peptide fragments from proteins separated by polyacrylamide gel electrophoresis. After completion of the electrophoresis step, gels are stained with Ponceau S or Coomassie Blue. Gel portions containing protein stained with Ponceau S are excised and transferred to borosilicate glass digestion tubes containing 0.9 ml of 1 mM NaOH or 5 mM Na2HPO4. After complete dissociation of the dye from the protein, 0.1 ml of 20% formic acid is added and the protein is hydrolyzed in situ at 112 degrees C for four hours. Subsequently the acid solution is made 10% in acetonitrile and chromatographed as such on a C18 (C4) reversed-phase column using an appropriate large-volume sample loading syringe and injection loop. Proteins stained with Coomassie Blue can be hydrolyzed in situ after complete removal of the dye with an aqueous solution containing 40% acetone, 10% triethylamine and 5% acetic acid. The gel slices are next washed with HPLC-grade water and protein is hydrolyzed in 2% formic acid under standard conditions. Gel-related contaminants do not interfere with the peptide separation under the proper conditions of HPLC analysis.     

A simple and inexpensive "cell dissociation sieve-tissue grinder" apparatus

A simple and inexpensive cell dissociation sieve-tissue grinder apparatus consisting essentially of stainless steel sieve (the one popularly used for sieving tea leaves) and a glass syringe plunger acting as pestle, is described for making single cell suspension.     

A method of removing gels from narrow-diameter glass tubes as part of a mini-two-dimensional electrophoresis procedure

A glass chamber that facilitates the removal of polyacrylamide gels from narrow glass tubes after the completion of isoelectric focusing is described. The tube is firmly held at one end of the glass chamber and a syringe is connected to the other end. By slowly forcing water from the syringe into the glass chamber, sufficient pressure can be generated to cause the rod gel to move slowly out of the glass tube. The rod gel is then used for the second stage of a mini-two-dimensional electrophoresis technique. Rod gels of 1.7-mm diameter have been used most extensively and can be removed from the glass tubes with almost 100% success.     

Development of a potentially wearable glucose sensor for patients with diabetes mellitus: design and in-vitro evaluation

A potentially wearable glucose sensor was developed, consisting of an oxygen electrode as detector and a dynamic enzyme perfusion system as selector. The selector is a hollow fibre, which can be placed subcutaneously and dialyses glucose from tissue fluid. In this design the problems of enzyme instability and oxygen limitation might be circumvented. The sensor measures glucose reliably for over two weeks, provided a new 10 ml syringe containing a glucose oxidase solution is connected to the system each day.     

An Apparatus for Measuring Volumes of Small Objects

An apparatus for measuring volumes of small objects such as tissue blocks is described. The apparatus measures volumes by fluid displacement and consists of a micropipette adapted to fit the mouth of an Erleiuneyer flask, a Luer adaptor fused to the side of the flask, and a glass syringe. When assembled with fluid enclosed, the fluid rises to a low level in the micropipette. Withdrawal of fluid into the syringe lowers the fluid level below the mouth of the flask. The micropipette is raised, the object to be measured is placed in the flask, and the micropipette is joined to the flask again. Fluid returned to the flask from the syringe rises to a higher level in the micropipette. The difference between the two fluid levels equals the volume of the object measured.

This apparatus gives reproducible measurements and can be calibrated for absolute volume determination. It is inexpensive to construct and easy to use.     

An Escherichia coli MutY mutant without the six-helix barrel domain is a dimer in solution and assembles cooperatively into multisubunit complexes with DNA

Escherichia coli MutY is an adenine and weak guanine DNA glycosylase involved in reducing the mutagenic effects of 7,8-dihydro-8-oxoguanine (GO). MutY contains three structural domains: an iron-sulfur module, a six-helix barrel module with the helix-hairpin-helix motif, and a C-terminal domain. Here, we demonstrate that the mutant MutY(Delta26-134), which lacks the six-helix barrel domain, cannot complement the mutator phenotype of a mutY mutant in vivo. However, the mutant can still bind DNA and has weak catalytic activity at high enzyme concentrations. The mutant is a dimer in solution and assembled into two and multiple (up to five) complexes with 20- and 44-bp DNA fragments, respectively, in a concentration-dependent manner. Higher order complexes with DNA substrates containing A/GO mismatches were formed at lower protein concentrations than with the A/G mismatch and homoduplex DNA. Measurement of equilibrium binding using fluorescence anisotropy showed that the mutant protein retains some specificity for A/GO-containing DNA substrates and that the binding event is highly cooperative. This is consistent with the MutY structure determined, which indicates that GO specificity is contributed by both the six-helix barrel and C-terminal domains. The nonspecific binding of MutY(Delta26-134) to DNA suggests a model in which the specific binding of mismatched DNA by MutY involves sequential interactions, in which one MutY molecule scans the DNA and enhances binding of another MutY molecule to the A/GO mismatch.