A Plexiglas Isolation-Transfer Chamber for Use with the Fonbrune Micromanipulator

The chamber is made with two pieces of clear Plexiglas, both 1 inch wide and 3 inches long; the top piece, 1/16 inch thick; the bottom, 1/4 inch. A recess 14 mm wide, 37 mm long and 5 mm deep is cut into the bottom piece leaving a margin 1.5 mm wide, this margin is then cut down to a height of 2.5 mm for the length of the recess; a piece of moistened filter paper I1/2 inches long and 5 mm deep is attached to the rear wall, leaving the bottom clear for the transmitted light; two notches 13 × 15 mm and 12 mm apart are cut from the top piece. The top is superimposed on the bottom and held by two screws to form a chamber that is accessible to microdissection instruments through its open side. Two cover glasses, one bearing the specimen and the other, the medium to which the transfer is to be made, are placed over the notches in the top (agar side down). The actual transfer of material is made by shifting the position of the cover glasses with the mechanical stage of the microscope while the specimen is held by the micromanipulator.     

A Staining Rack for Handling Cover-Glass Preparations

A porcelain staining rack (Fig. 1) has been devised for handling cover-glass preparations. The design is along the general lines of staining racks for slides commonly sold by commercial houses. It consists of three parallel rods—one at the bottom, two on the sides. These three rods are held together by two end pieces. Each of the rods has twelve slots, thus the rack holds twelve cover glasses. The slots have been arranged to hold the cover glasses some distance apart. Circular cover glasses having a diameter of 22 mm. are most desirable. Each of the two end pieces has a hole near the top for inserting one end of the wire tongs or handle (see Fig. 1) which is used for removing the rack from the stender dish. The staining rack is 40 mm. long and 35 mm. high and is designed, as shown in the figure, to fit the ordinary Stender dishes commonly used in many laboratories.     

A Staining Rack for Handling Cover-Glass Preparations

A porcelain staining rack (Fig. 1) has been devised for handling cover-glass preparations. The design is along the general lines of staining racks for slides commonly sold by commercial houses. It consists of three parallel rods—one at the bottom, two on the sides. These three rods are held together by two end pieces. Each of the rods has twelve slots, thus the rack holds twelve cover glasses. The slots have been arranged to hold the cover glasses some distance apart. Circular cover glasses having a diameter of 22 mm. are most desirable. Each of the two end pieces has a hole near the top for inserting one end of the wire tongs or handle (see Fig. 1) which is used for removing the rack from the stender dish. The staining rack is 40 mm. long and 35 mm. high and is designed, as shown in the figure, to fit the ordinary Stender dishes commonly used in many laboratories.     

Modification of the Carmen-Faull-Pullar Racks for Free-Floating Serial Sections

The original staining racks designed by Carmen et al. (Stain Techn., 43: 157-60) have been redesigned to use inside plates of 0.06 inch thickness and outside (top and bottom) ones of 3/16 inch. The greater thicknesses permit freer circulation about the sections and avoid the need for outer clamps to hold the assembly together. As in the original racks acrylic sheeting has been used, but with 25 × 45 mm holes for sections in each plate. Ordinary fiberglass window screening was cemented to one side of each plate. The assembly of 12 inner and the 2 outer plates was held together by 2 bolts made of 1/4 inch acrylic rod. Since clamps on the edges of the assembly were not needed, smaller staining dishes could be used, with coincident economy in volume of staining solutions.     

Apparatus for the Simultaneous Handling of Numerous Histological Specimens

An apparatus has been devised consisting of a series of bakelite plates 2 3/4 inches in diameter which can be clamped together one on top of the other. Each contains 37 depressions 1/4 inch in diameter, with a 1/16 inch hole at the bottom of each passing completely thru the plate. Each depression can contain its own specimen, its identity established by the number of the plate and the position of the depression. Several plates can be clamped together and run simultaneously thru the necessary reagents, thus handling large numbers of specimens without losing the identity of any of them.     

Integral Heating Device for the Rose Culture Chamber

The culture chamber consists of two metal plates held together by four short screws, a thin (film-type) electrical heating unit, a silicone rubber gasket and two cover slips. The order of assembly is bottom plate, heating element, first cover slip, gasket, second cover slip, and top plate. Syringe needles, one containing a thermistor, and others for supply and removal of fluid are inserted into the chamber through the gasket. Temperature is controlled by electrical connections through a Thermistemp temperature controller.     

A Holder for Simultaneous Fluid Processing or Carbon Coating of Electron Microscope Grids in Lots of 10 or More

A piece of polyethylene tubing about 14 cm long, ID 3 mm and with a 1.5 mm wall is shaped to hold grids in a linear series as follows. A series of pairs of transverse cuts I mm apart are made across and about $3/4 through the tubing at intervals of 1 cm. After making diagonal cuts on either side of each pair of cuts, wedge-shaped pieces of tubing are removed to leave free-standing 1 mm widths of the tubing wall. Each arch of wall is then bisected by a cut in the longitudinal direction of the tubing, thus providing the jaws of a clamp for a grid. By spreading the jaws with forceps, the grid can be inserted. On removal of the forceps, the grid is held firmly and when the holder is filled, the grids face in a direction perpendicular to its long dimension. By inserting a piece of glass tubing into the lumen of the holder, it is held straight and can be placed in a test tube for fluid processing. Stains or other fluids can be introduced through the glass tube, thus allowing fluid changes without exposure to air. Carbon coating can be achieved without removing the grids, since they all face the same way. For convenience in loading and unloading, the holder can be temporarily attached to a base with double-coated adhesive tape.     

A Desiccant-Holding Slide Box for Radioautographic Exposure

A double-depth plastic slide box is made by removing the bottoms of two standard 25-slide boxes and cementing the remaining frames together, bottom to bottom. A desiccant is placed in one box cover and fitted to the lower section of the cemented unit. A piece of screening is placed over the desiccant, the upper section filled with slides for radioautogaphy, and the empty cover applied to the top of the assembled box. Thus all slides are kept effectively and uniformly dehydrated during exposure.     

A Method of Eliminating the Electrification of Paraffin Ribbons

An apparatus for effectively eliminating the electrification of paraffin ribbons can be constructed for about $2. The essential parts are: a Ford T-TT induction coil to which is connected two, 4-inch brass or copper strips. To the end of one strip is soldered a copper disc 1 1/2 inches in diameter, which is then covered with heavy tinfoil. To the second strip is soldered one-half of a similar disc and its surface is also covered with several layers of tinfoil. The tinfoil should extend 1/2 inch beyond the straight edge of the half circle copper plate. The foil extending is cut to present 15 or 20 pointed projections. The coil is supplied with current from a toy train transformer yielding 5 to 12 volts or 4 to 6 amps. When the coil is in operation the two discs should be moved apart to a distance just beyond which the spark fails to jump. The apparatus is so set that the two brass strips are parallel to and 1 or 2 inches above the microtome knife. A bell push-button may be inserted into the circuit so that the operation of the apparatus may be controlled by the foot, leaving the hands free to handle the ribbon.     

一种改进的粉虱成虫生物测定方法

生物测定是检测害虫抗药性的一项重要技术。利用100mL插口圆底聚丙烯离心管对现在应用较多的粉虱成虫生物测定方法——琼脂保湿浸叶法进行了改进。改进后的方法不影响粉虱成虫的持续取食,具有操作简单、结果重复性好及无需对成虫麻醉等优点。同时发现茄子叶片对于B型烟粉虱Bemisia tabaci(Gennadius)B-biotype和温室白粉虱Trialeurodes vaporariorum(Westwood)成虫均是一种非常适合的生测材料。利用该方法分3次独立测定了烯啶虫胺对B型烟粉虱和温室白粉虱混合日龄成虫的毒力,结果具有很好的重复性。     

Simple Methods for Mammalian Chromosomes

The ordinary Feulgen-squash technic, after acetic-alcohol fixation, provides a simple and reliable method of preparing many mammalian tissues for chromosome counts. Tissues best suited to the technic were the seminiferous tubules. Small pieces of tissue about 3-6 mm. long and 1-2 mm. wide were immersed in a freshly made fixative (composed of 3 parts absolute ethyl alcohol and 1 part glacial acetic acid) immediately after removal by biopsy or from a freshly killed animal. After fixation for 3-12 hours the tissue was stained by the standard Feulgen procedure after hydrolysis for 12 minutes in normal HCl at 60 °C A 1-2 mm. piece was then teased apart on a slide in 45% acetic acid with a pair of mounted needles, and the teased tissue was squashed between the slide and the cover slip. During squashing the pressure was applied by hand and was regulated so as to avoid any excessive scattering of the chromosomes. The preparations were made permanent by dehydrating and mounting in Euparal.     

Enumerating phytoplankton with an upright compound microscope using a modified settling chamber

A modified settling chamber is described which permits the use of an upright compound microscope for phytoplankton enumerations. The chamber is composed of a 75 × 51 mm rectangular or 70 mm round-glass microslide base with a 130 m thick piece of sheet styrene attached to the upper surface. A circular cell is cut into the styrene material making a 26 mm diameter chamber which is approximately 130–150 m deep. Settling procedures follow Utermöhl's technique, with the use of a 0.13–0.15 mm thick coverslip (50 × 45 mm) to cover the chamber. The overall thickness of the settling slide is 1.13 mm which does not impact on the optical requirements for most objectives, including oil immersion objectives. The chamber encloses a total volume of 69–80 mm³. No statistical differences are observed in cell, filament or colony counts between the new and traditional chambers. Furthermore, count comparisons at different cell concentrations using the new chambers give consistent results. Thus, the resolution and availability of an upright microscope makes the use of the new settling chamber an attractive method for phytoplankton counting, especially in teaching situations.     

Production of Carbon Films for Electron Microscopy; Hydrofluoric Acid Detachment from Glass

Chemically clean microscope slides are coated as usual by vaporized carbon. The carbon film is floated off the slide by slowly lowering it at an angle of 45° into 1% HF in distilled water containing 0.025% Tween 80. This solution fills completely (forming a positive meniscus at the edges) one chamber of a double-compartment Perspex trough; the other compartment being similarly filled with the Tween solution only. A Teflon bar, laid on top of the partition keeps the solutions from mixing. After the carbon film loosens, it is floated across the central partition into the second compartment with the aid of a second Teflon bar, using both bars to guide the film on the surface of the fluid. The HF is thus washed from the film. Grids are thinly coated with 0.5% poly isobutylene in toluene (as an adhesive) and previously placed on a rectangle of filter paper supported by wire screening about 1/2 inch from the bottom of the trough. While the Tween solution is drained away through a bottom opening, the carbon film is guided to cover the grids. The filter paper bearing the grids is then removed and caused to dry slowly (about 12-16 hr) to avoid cracking or distortion of the film.     

Growth of Staphylococcus aureus MF 31 on the Top and Cut Surfaces of Southern Custard Pies

A Staphylococcus strain was inoculated on the top and cut surfaces of freshly baked Southern custard pies which were then packaged in a pasteboard carton and held at 30 C. Daily plate counts of surface sections 0.3 inch (0.76 cm) in thickness were made. The top surface inoculum showed a 24-hr lag time. This was due to the protective action of a top cakelike layer as shown by homogenization of the mix and coating of the surface. Substitution of all sweeteners with dextrose completely inhibited growth on the top surface. Further addition of dextrose to lower water activity (Aw) to 0.9 prevented growth on the cut surface as well, but such pies were organoleptically unacceptable. Growth on the top surface could also be prevented by 80 mug of undissociated sorbic acid per g in combination with 100 mug of undissociated propionic acid per g in the baked pie. Growth on the cakelike top surface was always retarded longer than on the cut surface provided the packaging allowed evaporation of surface moisture. Reducing the Aw of a different type of cream pie to 0.907 prevented top surface growth. It was concluded that baked cream pies with a cakelike top layer could be marketed with a "refrigerate after opening" label, provided the package maintains the moisture gradient caused by the surface skin and either a combination of 80 mug of undissociated sorbic acid per g and 100 mug undissociated propionic acid per g is present in the baked pie or the Aw of the baked pie is 0.920 or lower.     

Time‐lapse imaging system with shell‐less culture chamber

We have developed a system for imaging whole chick embryos from embryonic day 1.5 (E1.5) to E4.5. Our system consists of a custom‐made culture chamber, the top and bottom of which were heated and the inside was humidified. The system also has a fixed stage uplight fluorescent microscope, and long‐working distance objective lenses were adopted. The albumen removed‐yolk with the embryo in the dish was put in the chamber. It is of importance that we adopted long working distance lenses because the working distance of conventional objective lenses is too short for observation of the embryo in a humidified chamber. The objective lens we adopted has sufficient resolution to detect fluorescent protein expression at the single‐cell level. Transparent glass heater set on the top of the chamber helps to reduce dew condensation; the bottom heater keeps the temperature inside the chamber, and the water bath surrounding the egg maintains humidity. This system was used to detect fluorescent protein expressing cells in embryos. We could successfully trace those cells for 17 h in vivo. In conclusion, this system is useful for time‐lapse analysis of fluorescent protein expression and distribution for a longer period of time.     

Engineering a chemical implementation device and an imaging device for detecting chemiluminescence with a Polaroid™ high‐speed detector film: application to influenza diagnostics with the ZstatFlu®‐II test

We describe the engineering and product development of the chemiluminescent ZstatFlu®‐II Test kit for influenza diagnostics. The reaction vessel is a chemical implementation device with a polystyrene bottom chamber and a polypropylene top chamber that screw together. The patient's specimen is dispersed in a proprietary diluent and mixed inside the bottom chamber with the influenza viral neuraminidase‐specific substrate, 1,2‐dioxetane‐4,7‐dimethoxy‐Neu5Ac. Neuraminidase catalysis releases the dioxetane. The top chamber contains 40% NaOH and is sealed at the top with an ABS plastic plug‐crush pin assembly. The top chamber floor is 85% thinner at the centre, forming a frangible flap. An automated imaging device serves as an incubator for the chemical implementation devices and also facilitates the piercing of the flap by the crush pin. This action results in NaOH flushing into the bottom chamber, initiating chemiluminescence. The imaging device also exposes the Polaroid? high‐speed detector film to chemiluminescence. At the end of exposure, the film is automatically processed and ejected. Chemiluminescence from an influenza virus‐positive specimen produces a ‘+’‐shaped white image, archiving the diagnostic outcome. The modular ZstatFlu®‐II test kit components are easily adaptable for the chemiluminescent detection of a wide range of analytes. Copyright © 2003 John Wiley & Sons, Ltd.     

Climate‐mediated changes in marine ecosystem regulation during El Niño

The degree to which ecosystems are regulated through bottom‐up, top‐down, or direct physical processes represents a long‐standing issue in ecology, with important consequences for resource management and conservation. In marine ecosystems, the role of bottom‐up and top‐down forcing has been shown to vary over spatio‐temporal scales, often linked to highly variable and heterogeneously distributed environmental conditions. Ecosystem dynamics in the Northeast Pacific have been suggested to be predominately bottom‐up regulated. However, it remains unknown to what extent top‐down regulation occurs, or whether the relative importance of bottom‐up and top‐down forcing may shift in response to climate change. In this study, we investigate the effects and relative importance of bottom‐up, top‐down, and physical forcing during changing climate conditions on ecosystem regulation in the Southern California Current System (SCCS) using a generalized food web model. This statistical approach is based on nonlinear threshold models and a long‐term data set (~60 years) covering multiple trophic levels from phytoplankton to predatory fish. We found bottom‐up control to be the primary mode of ecosystem regulation. However, our results also demonstrate an alternative mode of regulation represented by interacting bottom‐up and top‐down forcing, analogous to wasp‐waist dynamics, but occurring across multiple trophic levels and only during periods of reduced bottom‐up forcing (i.e., weak upwelling, low nutrient concentrations, and primary production). The shifts in ecosystem regulation are caused by changes in ocean‐atmosphere forcing and triggered by highly variable climate conditions associated with El Niño. Furthermore, we show that biota respond differently to major El Niño events during positive or negative phases of the Pacific Decadal Oscillation (PDO), as well as highlight potential concerns for marine and fisheries management by demonstrating increased sensitivity of pelagic fish to exploitation during El Niño.     

一种适合于烟粉虱产卵的新型人工膜系统

介绍一种适合烟粉虱Bemisia tabaci(Gennadius)产卵的新型人工膜系统。该系统由一个透明的塑料盒,底直径30mm,顶口直径40mm,深35mm,其上覆盖1层或2层Parafilm M膜,与产卵的成虫相接触的1层厚度为5~8μm,2层膜中间或单层膜上加试验所需的试剂。将膜装置放于不透光的容器上,其上覆盖黄色玻璃纸,以吸引烟粉虱产卵。该装置可用于烟粉虱的人工产卵,观察卵的发育及研究相关的生理机制。     

Evolving strategies for making physical maps of mammalian chromosomes

Two types of physical maps are described: restriction maps made by top down approaches using enzymes that cut the genome infrequently, and complete libraries, made by bottom up approaches using fingerprinting of randomly selected cloned DNA. Construction of such maps for mammalian chromosomes is complicated by the mosaic nature of mammalian genomes, and extensive polymorphisms at the cleavage sites of most enzymes that yield large DNA fragments. However, it appears that both of these potential difficulties can be turned into advantages by new mapping strategies. When combined with yeast artificial chromosome cloning and polymerase chain reaction amplification methods, these approaches should soon yield complete maps of many human chromosomes.     

Analysis of the nest environment of tuatara Sphenodon punctatus

Water potential and temperature were monitored in 20 natural nests of tuatara, Sphenodon punctatus , through 12 months of incubation on Stephens Island, New Zealand. Tuatara nest in rookeries in open pasture, in sites that often are more than 100m from residential burrows located beneath the closed canopy of native bush. Nest tunnels are approximately 197 mm long, 73mm wide, and 45mm high, and have a slightly expanded chamber at the end. Eggs are generally deposited in 1-3 layers in the terminal chamber. The top eggs are 30-155mm below the soil surface, and an air space of as much as 20 mm may exist between the uppermost egg and the top of the chamber. Each nest receives an average of 8.6 eggs that imbibe water and swell during incubation. Only 48% of eggs have hatched or are still alive 12 months after oviposition. Survival by embryos is higher in moist nests than in dry ones. Variation in temperature in nests has only a small influence on survival, and this influence may be mediated indirectly by effects of temperature on the water exchanges experienced by incubating eggs. Water potentials in the soil of closed canopy forest on Stephens Island are high enough to support embryogenesis, but temperatures are too low. Thus, females leave the forest to nest in areas where soil temperatures are suitable for incubation.