Although the majority of microorganisms used in this laboratory are nonpathogens, some pathogens will be encountered.
It is for this reason that we must treat all accidental biological spills as if pathogens were involved. Chemical spills are just as important to report because some agents used in this laboratory may be carcinogenic; others are poisonous; and some can cause dermal damage such as blistering and depigmentation. Decontamination Procedure Once your instructor is notified of an accidental spill, the following steps will take place: 1.
Any clothing that is contaminated should be placed in an autoclavable plastic bag and autoclaved. Additional germicide should be poured around the edges of the spill to prevent further aerosolization. After approximately 20 minutes, the paper towels should be scraped up off the floor with an autoclavable squeegee into an autoclavable dust pan.
The contents of the dust pan are transferred to an autoclavable plastic bag, which may itself be placed in a stainless steel bucket or pan for transport to an autoclave. All materials, including the squeegee and dustpan, are autoclaved. Make it a habit to keep your hands away from your mouth. Obviously, labels are never moistened with the tongue; use tap water or self-adhesive labels instead.
Laboratory Protocol x.
Always clean up after yourself. Gram-stained slides that have no further use to you should be washed and dried and returned to a slide box.
Microbiological Applications: Short Version: A Laboratory Manual in General Microbiology
Coverslips should be cleaned, dried, and returned. Staining trays should be rinsed out and returned to their storage place. Return all bulk reagent bottles to places of storage. Return inoculating loops and needles to your storage container. Be sure that they are not upside down. If you have borrowed something from someone, return it.
Do not leave any items on your desk at the end of the period.
Do not disturb another class at any time. Wait until the class is dismissed.
Environmental Monitoring and Characterization
Treat all instruments, especially microscopes, with extreme care. Work cooperatively with other students in groupassigned experiments, but do your own analyses of experimental results. Laboratory Protocol xi. If you have had extensive exposure to microscopy in previous courses, this unit may not be of great value to you; however, if the study of microorganisms is a new field of study for you, there is a great deal of information that you need to acquire about the proper use of these instruments. Microscopes in a college laboratory represent a considerable investment and require special care to prevent damage to the lenses and mechanicals.
The fact that a laboratory microscope may be used by several different individuals during the day and moved around from one place to another results in a much greater chance for damage and wear to occur than if the instrument were used by only one individual. The complexity of some of the more expensive microscopes also requires that certain adjustments be made periodically. Knowing how to make these adjustments to get the equipment to perform properly is very important.
An attempt is made in the five exercises of this unit to provide the necessary assistance in getting the most out of the equipment.
Microscopy should be as fascinating to the beginner as it is to the professional of long standing; however, only with intelligent understanding can the beginner approach the achievement that occurs with years of experience. Microscopy 1 PART. Microscopy 1. At the end of each lab session be sure to wipe any immersion oil off the immersion lens if it has been used. More specifics about lens care are provided on page 5. Dust Protection In most laboratories dustcovers are used to protect the instruments during storage. If one is available, place it over the microscope at the end of the period.
Framework All microscopes have a basic frame structure, which includes the arm and base. To this framework all other parts are attached. On many of the older microscopes the base is not rigidly attached to the arm as is the case in figure 1. Stage The horizontal platform that supports the microscope slide is called the stage. Note that it has a clamping device, the mechanical stage, which is used for holding and moving the slide around on the A microscope that allows light rays to pass directly through to the eye without being deflected by an intervening opaque plate in the condenser is called a brightfield microscope.
This is the conventional type of instrument encountered by students in beginning courses in biology; it is also the first type to be used in this laboratory. All brightfield microscopes have certain things in common, yet they differ somewhat in mechanical operation. An attempt will be made in this exercise to point out the similarities and differences of various makes so that you will know how to use the instrument that is available to you. Before attending the first laboratory session in which the microscope will be used, read over this exercise and answer all the questions on the Laboratory Report.
Your instructor may require that the Laboratory Report be handed in prior to doing any laboratory work. The following suggestions cover most hazards. Transport When carrying your microscope from one part of the room to another, use both hands when holding the instrument, as illustrated in figure 1. If it is carried with only one hand and allowed to dangle at your side, there is always the danger of collision with furniture or some other object. And, incidentally, under no circumstances should one attempt to carry two microscopes at one time.
Clutter Keep your workstation uncluttered while doing microscopy. Keep unnecessary books, lunches, and other unneeded objects away from your work area. A clear work area promotes efficiency and results in fewer accidents. Electric Cord Microscopes have been known to tumble off of tabletops when students have entangled a foot in a dangling electric cord. Figure 1. Note, also, the location of the mechanical stage control in figure 1.
Light Source In the base of most microscopes is positioned some kind of light source. Ideally, the lamp should have a voltage control to vary the intensity of light. The microscope in figure 1. The microscope base in figure 1.
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Most microscopes have some provision for reducing light intensity with a neutral density filter. Such a filter is often needed to reduce the intensity of light below the lower limit allowed by the voltage control. On microscopes such as the Olympus CH-2, one can simply place a neutral density filter over the light source in the base. On some microscopes a filter is built into the base. Lens Systems All microscopes have three lens systems: the oculars, the objectives, and the condenser. Although the microscope in figure 1. Three or more objectives are usually present. Note that they are attached to a rotatable nosepiece, which makes it possible to move them into position over a slide.
The third lens system is the condenser, which is located under the stage. It collects and directs the light from the lamp to the slide being studied.
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The condenser can be moved up and down by a knob under the stage. A diaphragm within the condenser regulates the amount of light that reaches the slide. Microscopes that lack a voltage control on the light source rely entirely on the diaphragm for controlling light intensity. On the Olympus microscope in figure 1.
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On some microscopes a diaphragm lever is present. Focusing Knobs The concentrically arranged coarse adjustment and fine adjustment knobs on the side of the microscope are used for bringing objects into focus when studying an object on a slide. On some microscopes these knobs are not positioned concentrically as shown here.
Ocular Adjustments On binocular microscopes one must be able to change the distance between the oculars and to make diopter changes for eye differences. On most microscopes the interocular distance is changed by simply pulling apart or pushing together the oculars.
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To make diopter adjustments, one focuses first with the right eye only. Without touching the focusing knobs, diopter adjustments are then made on the left eye by turning the knurled diopter adjustment ring figure 1. One should now be able to see sharp images with both eyes. The optimum resolution of the best microscopes with oil immersion lenses is around 0.
This means that two small objects that are 0.