Microscopes PDF Study Notes

Study Notes on Care and Use of the Compound Microscope Initial Setup and Focusing To begin using a compound microscope, look into the eyepiece and adjust the coarse adjustment knob to raise the objective lens until the lines of the millimeter scale come into sharp focus. This is crucial for obtain...

Study Notes on Care and Use of the Compound Microscope Initial Setup and Focusing To begin using a compound microscope, look into the eyepiece and adjust the coarse adjustment knob to raise the objective lens until the lines of the millimeter scale come into sharp focus. This is crucial for obtaining a clear image of the specimen. Light Adjustment Adjust the light intensity by moving the iris diaphragm lever. Aim for a bright but comfortable illumination level to avoid over-illuminating the field, which can cause transparent objects to disappear. The fine adjustment knob should be used to focus on the location where three colored threads cross, determining their order from top to bottom based on which color is in focus at different depths. Measuring Depth of Field Position the millimeter ruler so that its scale crosses the greatest diameter of the field of view. Move the ruler so that one of the millimeter marks aligns with the edge of the field of view. The vertical depth of the specimen that is clearly in focus is referred to as the depth of field. Continue using the fine adjustment knob to determine relative depths of structures within the specimen. Purfocus Feature Most microscopes are designed to be parfocal, meaning that when a specimen is in focus with a lower-power objective, it will remain in focus (or nearly so) when a higher-power objective is rotated into position. Always center the specimen in the field of view before changing to higher objectives. Using the Micrometer Scale Some compound microscopes come equipped with a micrometer scale in the eyepiece, which is subdivided into fifty to one hundred equal divisions. These divisions can be calibrated against known divisions of a micrometer slide placed on the microscope stage. This allows for the measurement of the length and width of microscopic objects by superimposing the scale over the magnified image. High-Power Objective Focusing When rotating the high-power objective into position, focus using only the fine adjustment knob. Using the coarse adjustment knob at high power can accidentally force the objective into the coverslip due to the shorter working distance. Adjust the iris diaphragm for proper illumination, as more light is typically needed at higher magnifications. Preparing Wet Mounts To prepare several temporary wet mounts using small, transparent objects, follow these steps: 1. Carefully clean a microscope slide with soap and water, then dry it with a paper towel. 2. Place a tiny, thin piece of the specimen in the center of the slide. 3. Use a medicine dropper to add a drop of water over the specimen 4. Consult with your instructor if a drop of stain might enhance the image of any cellular structures. Final Steps After preparing the wet mount, examine the specimens first using the low-power objective and then switch to the high-power objective to observe their details. Always ensure proper focus and illumination throughout the process to achieve the best Care and Use of the Compound Microscope Microscope Objectives The compound microscope typically has multiple objective lenses, with common powers being 40x and 100x. The power of each objective is indicated on the metal casing. To prevent the objective from touching the specimen, observe from the side while focusing. Total Magnification Calculation To calculate the total magnification, multiply the power of the eyepiece by the power of the objective lens. For example, using a 10x eyepiece with a 40x objective results in a total magnification of 400x. Microscope Care Rules It is essential to handle the microscope with care due to its delicate nature. Here are some key rules for proper care: Always hold the microscope by its arm with one hand and support the base with the other when moving it. Keep the microscope covered with a dust cover and stored in a cabinet when not in use. Clean the lenses gently with lens paper or a high-quality cotton swab. For additional cleaning, follow specific lens cleaning techniques. Ensure that the electric cord of the substage lamp does not hang off the table to prevent tripping hazards. Allow the lamp to cool before moving the microscope to extend bulb life. Never attempt to disassemble the microscope or remove its parts. If the microscope is malfunctioning, report the issue to your laboratory instructor immediately. Measuring Field of View To measure the diameter of the field of view, focus the lenses on a millimeter scale using a transparent plastic ruler. Follow these steps:  1. Place the ruler on the microscope stage using the spring clamp or under the stage clips. 2. Center the millimeter scale in the light beam from the condenser and rotate the scanning objective into position. 3. While observing from the side, lower the objective close to the ruler using the coarse adjustment knob, then refine the focus with the fine adjustment knob. 4. Turn on the substage illuminator and look through the eyepiece to see the illuminated field of view. Lens Cleaning Technique Proper lens cleaning is crucial for maintaining microscope performance. Follow these steps for effective cleaning: 1. Moisten one end of a high-quality cotton swab with a drop of Kodak lens cleaner, keeping the other end dry. 2. Clean the optical surface using the wet end in a circular motion, then dry with the dry end. 3. Use a hand aspirator to remove any lingering dust particles. 4. Start cleaning with the scanning objective and progress to higher magnifications, using a new cotton swab for each objective. 5. When cleaning the eyepiece, avoid opening the lens unless absolutely necessary. 6. For difficult cleaning, use alcohol as a last resort, and avoid regular use of xylene as it can damage lens coatings Care and Use of the Compound Microscope Introduction to Compound Microscopes Compound microscopes are essential tools in various scientific fields, allowing for the magnification of small specimens. They typically feature a monocular or binocular body and a mechanical stage for specimen manipulation. Major Parts of a Compound Microscope The compound microscope consists of several key components, each serving a specific function:  Eyepiece (Ocular): The lens through which the viewer looks to see the magnified image. Body Tube: Connects the eyepiece to the objective lenses, ensuring proper alignment. Body: The main structure that houses the optical components. Arm: Supports the body and connects it to the base. Nosepiece: Holds the objective lenses and allows for rotation to switch between them. Objectives: Lenses with varying magnification powers, typically marked with their respective magnification levels. Mechanical Stage: A platform that holds the slide and allows for precise movement. Stage Control Knobs: Used to move the mechanical stage in different directions. Coarse Adjustment Knob: For making large adjustments to the focus. Fine Adjustment Knob: For making small, precise adjustments to the focus. Condenser: Focuses light onto the specimen. Iris Diaphragm Lever: Controls the amount of light reaching the specimen. Substage Illuminator (Lamp): Provides the necessary light for viewing the specimen. Illuminator Switch: Turns the light source on and off. Base: The bottom support structure of the microscope. Understanding Magnification and Numerical Aperture Magnification is a crucial aspect of microscopy. The power of the eyepiece and objective lenses is indicated on the microscope. For example, a 10x eyepiece combined with a 40x objective provides a total magnification of 400x. The numerical aperture (NA) is a critical specification for objective lenses, indicating their light-gathering ability. For instance, a 40x objective with a numerical aperture of 0.65 can gather a significant amount of light, enhancing image clarity and detail. Conclusion Understanding the components and functions of a compound microscope is essential for effective use in scientific research. Proper care and handling of these instruments will ensure longevity and optimal performance Materials Needed To conduct observations using a compound microscope, the following materials are required: Compound microscope Lens paper Microscope slides Coverslips Transparent plastic millimeter ruler Slide of three colored threads Medicine dropper Dissecting needle (needle probe) Specimen examples for wet mounts Methylene blue (dilute) or Bodine-potassium-iodide stain Micrometer scale Stereo microscope (dissecting microscope) Procedure Begin by observing the compound microscope and studying its major parts as illustrated in the provided figure. The lens system of a compound microscope consists of three main components: Condenser Objective lens Eyepiece Light enters the microscope system from a substage illuminator (lamp) or mirror, which is then concentrated and focused by the condenser onto the microscope slide or specimen placed on the stage. The condenser is typically kept in its highest position to optimize light concentration. The iris diaphragm, located between the light source and the condenser, allows for the adjustment of light intensity. To observe the effect of the diaphragm, locate the lever beneath the stage and move it back and forth to see how it changes the size of the opening. Some microscopes may have a disc diaphragm instead, which has various sized holes for light admission. Understanding Microscope Functionality The compound microscope is essential for studying small structures, such as cells, as the human eye cannot perceive objects smaller than 0.1 mm in diameter. The compound microscope utilizes two sets of lenses: the eyepiece lens system and the objective lens system. The objective lens system magnifies the image, which is then further magnified by the eyepiece lens system, allowing for a total magnification of up to one thousand times. After light passes through a specimen on a microscope slide, it enters the objective lens system, projecting the light upward into the body tube to create a magnified image. The eyepiece lens system then magnifies this image further, typically at a power of 10x. Check the eyepiece for its power rating to confirm this. Purpose of the Exercise The primary goal of this exercise is to familiarize oneself with the major parts of a compound microscope and their functions, as well as to effectively use the microscope to observe small objects. Learning Objectives Upon completing this exercise, you should be able to: 1. Locate and identify the major parts of a compound microscope. 2. Describe the functions of these parts. 3. Calculate the total magnification produced by various combinations of eyepiece and objective lenses. 4. Prepare a simple microscope slide. 5. Make proper use of the microscope to observe small objects. Additional Notes Common objective lenses include: Scanning objective (4x) Low-power objective (10x)  High-power objective (high-dry-power) Understanding how to switch between these lenses using the revolving nosepiece is crucial for achieving different magnifications during observations To increase the light intensity, the rheostat is turned clockwise, and the iris diaphragm opening is progressively enlarged. The condenser will concentrate the light beam onto the specimen; therefore, the light must be positioned in the center of the field of view.