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Instruments A. optical microscope: Uses Light rays, which change direction when they pass form one transparent medium to another. Glass lenses ben light rays. 1. Simple: One lens, magnifying glass 2. Compound: uses two lenses. The Objective lens magnifies the object first, and then the eyepiece magnifies the second image. Parts and their functions Mechanical system: base, arc, stage, clips, body tube, course adj, fine adj Optical system: Ocular &Objective (High &Low power Light: a light source (mirror/lamp}. Diaphragm (regulates the light) (disk/iris), condenser (concentrate the light). Parts of the microscope = Ocular X Total magnification: Objectives, 100x = (10x) X (10x) Resolution: ability to show two points as separate. Sharpness of an image. Optical scopes are 500x better tan the eye. Optical Microscope Diaphragm: regulates the light Stained: Only certain parts absorb the stain to see that part better. Can kill the cell. Stereoscope Dissection scope/binocular scope): for studying external structures. For opaque specimens. Magnifies 6x to 50x. Provides a 3D image. Has an ocular and objective for each eye. Phase contrast: Contrast among cell parts without stains. Electron Microscopes Electron microscopes: uses electrons, not light. Electromagnets act as lenses to focus the electrons. The more dense parts of the specimen don’t allow the electrons to pass. Advantages: increased magnification, increased resolution. Disadvantages: ultra thin specimens and can only use dead organisms (vacuum) TEM Transmission Electron Microscope Magnify up to 250,000x Resolution: 10,000 times greater then the eye. Specimen must be ultra thin. May stain with atoms of heavy metal, which attach to certain parts. SEM (Scanning Electron Microscope): Lower magnification. Surface detail. 3D. Electrons are shot at it. The specimen is covered with gold, which gets excited. The electrons that are excited are collected on a screen. Biological Techniques A. Fixing/Embedding/Staining 1. material for microscopic examination must be thin enough to let light pass through. Transparent. 2. These methods prepare larger samples (tissues/organs) for microscope examination. Fixing: hardens the specimen Embedding: put in paraffin Sectioning: sliced thin Microtome: does the slicing Centrifugation Separates materials based upon their differences in density (more dense will spin to the bottom. materials to be separated are put in a liquid in a test tube. Ultracentrifuge: high speed. Used for cell parts. Spins at 40,000 to 100,00 revolutions. Microdissection 1. This is a dissection usually performed on microscope specimen, such as cells. 2. Some equipment used: microscalpel, microprobe, microneedle, micromanipulator. Tissue culture: 1. Living issue is grown (Cultured) in a solution. 2. Outside of the body. Gets all the nutrients it needs. Used in medical research (Cancer, Aids) Chromatography 1. Chemical substances from a specimen are separated/isolated using a solvent (dissolver) 2. The degree of separation depends upon the solubility of the materials in the mixture and relative adherence to the paper used. Electrophoresis 1. Material from a specimen are separated based upon their electrical charge