THE USE OF THE BRIGHTFIELD COMPOUND MICROSCOPE Flashcards
is a branch of science that deals with the study of bacteria.
Bacteriology
are minute organisms, most of which are too small to be seen with the naked eye.
Bacteria
The word_____ means that a specimen positioned properly on the stage of a microscope and illuminated by a light source will be magnified by a two-lens system.
compound
The image formed by the____ lens will be magnified again by the____ lens system. Therefore, the magnification is compounded.
objective
ocular
A common example of a compound microscope is the_______
The term______ indicates that magnified objects appear dark against a bright background.
A sufficient_____ must exist between the magnified object and the background for the objects to be visible.
Brightfield compound microscope.
brightfield
contrast
The magnification is achieved when light rays from a light source passes through a________, which has lenses that direct the light rays through the specimen.
condenser
From the condenser…Light rays then pass into the____, the lenses closest to the specimen.
Then the image of the specimen is magnified again by the______
objective lenses
ocular lens, or eyepiece
To preserve the direction of light rays at the highest magnification,______ is placed between the glass slides and the ______ objective lens.
immersion oil
OIO
Why is immersion oil used?
The immersion oil has the same refractive index as the glass slide, so the oil becomes part of the optics of the glass.
Why is it important that the refractive index of oil is same as the glass slide?
When immersion oil is used, the light rays do not refract when as they enter the air from the slide.
Therefore, the use of immersion oil improves the resolving power of the lenses.
If oil is not used with an oil immersion objective lens, the image becomes fuzzy, with poor resolution.
A compound brightfield microscope consists of (2) components.
structural and optical
Structural component
Head/ body
Base
Arm
Nose piece
Coarse adjustment knob
Fine adjustment knob
Stage
Stage clips
Illuminator
Condenser
Iris diaphragm
Condenser focus knob
Optical components
Eyepiece/ ocular
Eyepiece tube
Objective lenses
- houses the optical parts in the upper part of the microscope
Head/Body
supports the microscope and houses the illuminator
Base
connects to the base and supports the microscope head.
It is also used to carry the microscope.
Arm
” remagnifies the image formed by the objective lens.
Eyepiece or Ocular
Typically, standard eyepieces have a magnifying power of
10x
” holds the eyepieces in place above the objective lens.
Eyepiece tube
• primary lenses that magnifies the specimen.
Objective Lenses
” often marked with 4X.
Scanner
• often marked with 10X
Low Power Objective (LPO)
• It is usually shorter than the other two objectives
• It usually forms the general outline or wider portion of the object.
Low Power Objective (LPO)
• often marked with 45%
High Power Objective (HPO)
• It is longer than the LPO.
• It forms a bigger image of the object in focus.
In most cases it is used to enlarge specimens that are so small under LPO.
HPO
” often marked with 100X.
Oil Immersion Objective (OIO)
It has the highest degree of magnification and is used to examine stained smear preparations of microorganisms using_____ as their medium.
OIO
immersion oil
• a rotating turret that houses the objectives.
Nosepiece
• a bigger wheel used to adjust the low power objective in focusing.
Coarse Adjustment Knob
It is also used for initial focusing of the specimen.
Coarse AK
a smaller wheel used for delicate and final focusing of the specimen
when using the high power and oil immersion objectives.
Fine Adjustment Knob
• It is also used to make the specimen more vivid.
Fine AK
a square platform with an opening at the center.
Stage
It is where the slide/specimen is placed during focusing.
Stage
paired structures found on either side of the stage.
It is used hold the slide in place.
Stage Clips
the light source for a microscope, typically located at the base of the microscope.
Illuminator
” a lens found beneath the hole of the stage.
Condenser
It is used to collect and focus the light from the illuminator on to the specimen.
Condenser
It controls the amount of light reaching the specimen.
Iris Diaphragm
located above the condenser and below the stage.
Iris Diaphragm
• moves the condenser up or down to control the lighting focus on the specimen.
Condenser Focus Knob
- Get the microscope from the stockroom by taking the____ with one hand and supporting the instrument at the____with the other hand.
arm
base
- Place the microscope at least_____ from the edge of the laboratory table with the microscope____ facing you.
six inches
arm
- Wipe all lenses with ____before and after use.
If the oil-immersion lens is sticky, moisten a piece of lens paper with _____to wipe it clean.
If it is still sticky, wipe the lens clean with____, and immediately remove with ______then wipe dry the lens with____.
Consistent use of____ may loosen the lens.
Lens paper
95% alcohol
xylol
tissue moistened with 95% alcohol
lens paper
xylol
- Never use_____ when cleaning the objective lenses or oculars.
Failure to clean the lens will result in the formation of gummy residue which reduces the efficiency of the lens.
tissue or cloth
- Keep the stage..
clean and dry
- Do not____ the microscope; instead, adjust your____ so you can comfortably use the instrument.
___tends to distort wet mounts or objects covered with oil.
tilt
stool
Tilting
CORRECT MANIPULATION OF THE BRIGHTFIELD COMPOUND
MICROSCOPE
- Place the specimen slide on the stage and secure it with the stage clips.
Arrange the portion of the slide to be examined over the central opening in the stage. - Rotate the low-power objective into place under the body tube. You will feel a click when it is correctly in place.
- Raise the condenser as high as it will go.
- Rotate the coarse adjustment clockwise to bring down the low power objective (LPO) close to but not touching the slide, until the specimen is seen through the eyepiece.
- Regulate the intensity of light by opening or closing the iris diaphragm.
- Sharpen the focus by turning the fine adjustment knob.
- Turn the low power objective to high power objective without elevating the body tube. Notice that it will be almost in focus because most microscopes are parfocal. Little adjustments with fine adjustment knob will only be needed to clearly view the object in focus.
- To focus the oil immersion lens, swing the oil immersion objective halfway towards the specimen in focus. Place a drop of immersion oil on the smear.
Swing the oil immersion lens in place. Using the coarse adjustment, bring the objective down until it touches the oil. Find the object by gently turning the coarse adjustment knob upwards. Sharpen the focus with the fine adjustment knob. - Compare the relative sizes from those seen under the low power, high power and oil immersion objectives.
- Clean the stage and the lenses after each use, before returning the microscope to the stockroom.
CORRECT MANIPULATION OF THE BRIGHTFIELD COMPOUND
MICROSCOPE: summary
- Specimen on stage
- LPO
- Condenser- position high
- Coarse AK (cc)
- Iris diaphragm
- Fine AK
- LPO to HPO
- OIO + immersion oil. Adjust Coarse AK, touch oil. Fine AK
- Compare the sizes
- Clean stage & lenses
CALCULATING THE TOTAL MAGNIFICATION
The number tells the operator how many times the focused specimen is magnified. This is computed as follows:
Magnifying power of the eyepiece
X
Magnifying power of the objective
=
Total Magnification
The specimen in focus is magnified____ times its actual size using the oil immersion objective.
1000
____ has a smaller aperture than the ____, hence it will be necessary to open the ______ further in order to fill the objective aperture with light
HPO
LPO
iris diaphragm
The________ between the objective and the specimen slide differs according to the objective being used.
working distance (WD)
The working distance when using a scanner is______.
18.5 mm
In low power objective it has distance of_______.
10.6 mm
A distance_____ is observed in high power objective
0.6 mm
Working distance for oil immersion objective is______
0.13 mm
The last two working distances (HPO and OIO) are especially short; hence ALWAYS focus______ from the slide and NEVER focus______ on a slide while looking into the ocular to avoid damaging the objective lens.
AWAY
down
