Transcript USE AND CARE OF THE MICROSCOPE

USE AND CARE OF THE
MICROSCOPE
LECTURE 1
MICROSCOPY
 Light
Microscopy: any microscope that uses
visible light to observe specimens
 Compound Light Microscopy:
 utilizes
a series of lenses
 uses visible light as the source of illumination
 allows you to observe a specimen as a
magnified image
 Resolution=0.2 um
 Brightfield
Illumination: visualization of
dark objects in a bright field
Fine adjustment
Coarse adjustment
Key Terms
 Illuminator: light source
 Condenser: series of lenses that direct light through the
specimen of interest
 Objective lens: first magnification, lens located
closest to the specimen, usually 4 different objective lenses
(scanning, 10X, 40X, 100X)
 Ocular
lens
lens: located in the eyepiece, last magnification
 Coarse adjustment: used with low-power objectives
(scanning and 10X)
 Fine adjustment: used with high-power objectives
(40X and 100X)
Key Concepts
 Total
Magnification: Multiply the objective lens
magnification by the ocular lens (usually 10X) magnification
 Resolution/Resolving Power: Ability of the lens to
distinguish fine detail and structure or the ability to distinguish
two points a specified distance apart. Compound Microscope =
0.2 um. Shorter the wavelength, the greater the resolution.
 Oil Immersion: Same effect of increasing the objective lens,
therefore it improves the resolving power of the lens
 Refractive Index: The measure of the light bending ability of
a medium (Ex:Water). Therefore, specimens must be made to
contrast their medium.
 Use of Stains, Dyes
Other Types of Microscopy
 Darkfield: Specimen is light against a dark background, used to
examine live organisms
 Phase-Contrast: uses special condenser to enhance
intracellular details of specimen can be visualized in living
organisms/ takes advantage of differences in refractive indexes of
cell structures
 Differential Interference Contrast (DIC): Similar to
Phase-Contrast, higher resolution, image appears 3-D
 Fluorescence: Ability of specimens to absorb short
wavelengths of light (ultraviolet) and give off long wavelengths of
light (visible). Used in diagnostic procedures called fluorescent
antibody technique
 Confocal: specimen is stained w/ fluorescent dye, computer
process the image-uses laser light to illuminate one plane of a
specimen at a time. Higher resolution, clear images
Electron Microscopy
Electron Microscopy:
 Used for specimens smaller than 0.2 um
 Electrons used instead of a beam of light
 Higher Resolution due to shorter wavelengths
of electrons (~100,000X shorter than visible
light)
Types
of Electron Microscopes:
Transmission
Scanning
TEM
 Transmission
 A beam
Electron Microscopy (TEM):
of electrons is passed through an ultra-thin
section of specimen
 electromagnetic lenses are used to direct the beam
 a copper mesh grid is used to view the sample
 a projector lens is used to focus the image on a
fluorescent screen
 Resolving Power =2.5 nm
 Total Magnification =10,000-100,000X
 best for virus particles, flagella and proteins
 resulting photo is black and white, no 3-D
SEM
 Scanning
Uses
Electron Microscopy (SEM):
an electromagnetic lens to direct
electrons over the surface of a specimen
Results in a 3-D image
Resolving Power= 20 nm
Total Magnification = 1,000-10,000X
Best for observing the surfaces of
microorganisms
IMPORTANT TIPS TO
REMEMBER:
1-OCULAR AND OBJECTIVE LENSES
SHOULD ONLY BE CLEANED WITH LENS
PAPER (NOT BIBULOUS PAPER) ALWAYS
CLEAN THEM BEFORE AND AFTER USE
 2-OIL SHOULD ONLY BE USED WITH 100X
OBJECTIVE
 3-RETURN MICROSCOPES ON SCANNING
OBJECTIVE WITH STAGE UP AND SLIDE
CENTERED
 4-ALWAYS HANDLE AND CARRY YOUR
MICROSCOPE USING THE BASE AND ARM
