Section G Flashcards
compound microscope is composed of two lenses.
- a primary magnifying lens (ocular)-closest to the eyes and usually magnifies objects 10x their actual size
- a secondary lens system (objectives)-are a collection of lenses located in the nosepiece with different degree of magnification
Base
Supports the microscope and contains the light source, held by one hand when carrying microscope
Light Source
Located in the base.Provides illumination for the specimen, has a dimmer switch to control brightness
Arm
Supports the stage and body tube, held by other hand when carrying microscope
Condenser
Lens system below the stage (sub-stage) that focuses and concentrates light on the specimen . Has an adjustment knob, that raises and lowers the condenser to verify light delivery. Contains iris diaphragm.
Iris Diaphragm
Adjustable opening like iris of eye, regulates amount of light (diameter of light beam) passing through the condenser. Controlled by a lever, that permits the best possible contrast when viewing a specimen.
Course adjustment
Used to focus on the specimen. Rotating knob that rapidly brings the specimen into focus by raising /lowering the stage, used only with low power objective.
Fine adjustment
Rotating knob that slowly brings the specimen into sharp focus,used for critical focusing at low power and for all focusing at high power magnification.
Condenser control
Rotating knob that moves the condenser lens system up and down
- The condenser is very important in image formation.
- Like a microscope objective, a condenser has a numerical aperture and it should equal or better that of the highest magnification objective being used.
Mechanical stage
The platform that slides are placed on while being viewed. Platform has an opening in the center to allows light from lenses to pass through it as well as the specimen.
Usually has a spring clip to hold the slide in position for viewing.
Vernier scales
Set of scales on X and Y-axis that allow for the location of a particular object on a slide.
Stage control
Rotating knobs located under the stage that move the stage from left to right and back and forth.
Objectives lenses
Glass lens nearest the stage, magnifies image of the specimen by amount shown on the side of the lens producing a real, inverted image.
Lens system:10X (scanning lens), 40X (low power/high dry lens), and 100X(high power/oil immersion lens)
• The objective lens magnifies the specimen to produce a REAL IMAGE that is projected to the ocular.
Revolving nose piece
Rotating mechanism for objective lenses (usually 3-4) so they may be alternately positioned into the light path for different magnification.
Body tube
Joins nosepiece to oculars, has mirrors to pass image from objective lens to oculars.
Ocular
eye piece
One ot two glass lens closest to eye. Observations are made through the ocular(s).
An ocular lens has a magnification of 10X . (increases the size of the object by 10 times)
Magnifies real image from the objective lens, usually by producing a virtual, enlarged image.
Diopter
Focusing ring located on the ocular lens, allows for sharp focusing withboth eyes.
What is magnification
microscope is an instrument of magnification
- Magnification is the ability to increase the size of an image–not clarity
- Magnification uses lenses ( in the microscope the ocular lens and the objective lens)
resolution of a microscope
ability to clearly determine two close objects as separate or as singular, distinguished entities.
• The shortest distance between two points on a specimen that can still be distinguished by the observer
• The microscope has a resolution of 0.2 um
• The human eye can resolve objects about 100um apart
• If the objects are closer than 0.2 um, they will appear fused as a single object or points will blur together, making it impossible to differentiate
• If two objects are 0.01 mm apart, we cannot detect them unless we magnify an image of them by 10X
• Resolving power
is determined by the amount and physical properties of visisble light (wavelength spectrum of light ) entering the microscope.
• The more light delivered to the objectives the greater the resolution
• Blue light gives better resolution than yellow or red
• Blue filter used on microscope to “hold Back” longer wavelength of light
NUMERICAL APERTURE
The numerical aperture of a microscope objective is a measure of its ability to resolve fine specimen detail.
The numerical aperture of a lens is dependent upon two parameters
- The angle of the incidence of light onto the lens
- The refractive index of the glass of which the lens is composed.
- The size of the objective lens aperture (opening) decreases with increasing magnification, allowing less light to enter the objectives
When using the microscope it may be necessary to increase the light intensity as the magnification gets higher.
REFRACTIVE INDEX (R I)
ability of the microscope to both magnify and resolve (or allow small structures to be seen) depends on the refraction or bending of light
speed at which light passes through a medium of one density to another in a vacuum
Refraction
bending of light as it passes from one medium to another
- Immersion oil which has the same refractive index as glass is used to replace the air. The slide and lens are joined by a layer of oil
- As the wavelength of light increases, the refractive index decreases.
• Scanning power objective:
shortest objective. This lens magnifies objects four times actual size (4X).
• Low power objective:
The next shortest objective. This lens magnifies objects ten times their actual size (10X). Also used for scanning
Ocular x Objective = 10 x 10X = 100 Total magnification
• High-dry objective:
This is usually either the longest, or second longest objective. This lens typically magnifies objects forty times their actual size (40 X).
• Oil immersion objective:
The oil immersion lens magnifies objects one hundred times (100X), and must be used with a drop of oil placed directly on the specimen and fills the space between the lens and specimen.
Vernier Scales (Coordinates)
• The X coordinate is obtained using the scale at the top of the stage running from right to left
• The Y coordinate is obtained using the scale running from top to bottom along the side of the stage.
LEARN TO READ
Daily and weekly maintenence
- When not in use, keep the microscope protected with a plastic cover.
- Unplug microscope and wash hands.
- Accumulated dust in a microscope can deteriorate image quality. Keep all openings covered so that dust does not enter the microscope and settle on lenses, mirrors, and prisms.
- Wipe the stage with a damp cloth and mild detergent, and clean off any immersion oil with ethanol or lens cleaner.
- Carefully clean the objectives (see detailed description below).
- Remove and clean any dust or immersion oil from the condenser and turret using dry Kimwipe. Weekly give immersion lens a through cleaning (see description below)
- Remove the condenser top lens and clean it with lens paper and diluted methanol , if necessary.
- Remove both eyepieces and clean their surfaces with a Kimwipe and lens cleaner.
Objective Cleaning
- A through cleaning of immersion objectives is necessary at least once peer week. Some cleaning agents can erode the anti-reflection coating over time.
Remove the objective from the turret.
Add a few drops of diluted methanol to lens tissue.
Gently wipe the lens in a circular motion (only letting the tissue, not your fingers, come into contact with the lens element).
Remove any excess methanol immediately with a dry piece of tissue – allowing ethanol to remain on the lens could erode the anti-reflection coating.
For stubborn smears, place a drop of xylene or benzene on lens paper. Gently wipe the lens in a circular motion. Remove xylene or benzene with diluted methanol.
Xylene or benzene is toxic, so wear PPE and use under a fume hood
Köhler illumination
August Köhler of the Carl Zeiss corporation as a method of providing the optimum specimen illumination.
course adjustment knob to bring the stage and objective close together…bringing your object into focus. Watch the objective lens to make sure that the lens does not break the slide.
needs a high density illumination source, field diaphragm, condenser diaphragm, and collector and condenser lenses.
