2. tools in microbiology
TRANSCRIPT
Microscope instrument used to study MO Optical instrument used to observe tiny objects that
cannot be seen by the unaided human eye.
Simple Microscope - contains only one magnifying lens Anton van Leeuwenhoek
Compound Microscope - contains more than one magnifying lens. Compound light microscope. Hans Jansen and son Zacharias.
Photomicrographs - photographs taken through the lens system of compound microscopes.
PARTS OF A COMPOUND MICROSCOPE
Magnifying Parts:
-To enlarge objects of study
- objectives:
- Eyepiece or ocular objective: variable magnification
- Scanner: 5x magnification, used to study larger
organisms
- Low Power Objective (LPO): 10x magnification
- High Power Objective (HPO): 40x magnification
- Oil Immersion Objective: 100x magnification
Illuminating Parts:- Parts that modify light and illuminate object of
study- Abbe condenser: concentrates light- Mirror: reflects light or uses bulbs as main
light source- Iris diaphragm: regulates the amount of
light that hits the object of study Mechanical Parts:
- Supports the magnifying and illuminating parts- Used to focus the lenses
- Draw tube, body tube, revolving nosepiece, dust shield, arm, stage, stage clips, coarse adjustment knob, base, inclination joint
I. TYPES OF MICROSCOPE
A. VISIBLE LIGHT MICROSCOPY1. Bright- Field Microscope
---used to observe morphology of the organisms.
---does not resolve very small specimens (viruses)2. Dark-field Microscope
---”Dark” background, light organisms---used to detect Syphilis (Treponema
pallidum)
VISIBLE LIGHT MICROSCOPY
3. Phase-Contrast Microscope---Observe dense structures---To facilitate detailed
examination of the internal structures of living specimens.
4. Fluorescent Microscope
---Ultraviolet light
---used to show antibodies
B. ELECTRON MICROSCOPE
1. Transmission Electron Microscope
---Highest magnification (10,000- 100,000x)
---Cellular ultra structure and viruses
---2-D image
2. Scanning Electron Microscope
---Surface structure of cells and viruses
---3-D image
---magnification: (1000-10,000x)
Metric System
– used to describe sizes of microorganisms
1. decimeter 10
2. centimeter 100
3. millimeter 1000
4. micrometer 1M – bacteria, protozoa
5. nanometer 1B - viruses
II. STAINING PROCEDURESOBJECTIVES:
1. Kill the organism2. Preserve morphology3. Anchor smear to slide
A. Simple staining- Aqueous or alcohol solution of a single
basic dye- Used to determine size, shape and morphological arrangement- Ex. Methylene blue
Simple Staining Based on shape:
- Cocci: round or spherical bacteria
- Bacilli: rod-shaped or cigar-shaped bacteria
- Spirals: coiled bacteria
- Spirillum: flexible coiled bacteria
- Spirochetes: rigid, coiled bacteria Based on Arrangement of cells:
- Strepto: bacteria in chains
- Staphylo: bacteria in clusters
- Diplo: bacteria in pairs
- Tetra: bacteria in 4s
B. Differential staining – use of 2 or more dyes that may differentiate one type of organism from one another.
1. Gram Stain - used to classify MO- Dr. Hans Christian Gram (1884)
- Procedure: V I A S a. Crystal VViolet (primary stain) b. Gram’s Iodine (Mordant) c. Alcohol (Decolorizer) d. Safranin (Counterstain)
Differential staining
2. Acid-Fast Stain (Ziehl-Nielsen)- Binds strongly to the bacteria that
have a waxy material in their cell wall- Used to identify Mycobacterium, Nocardia
- Procedure: C A M a. Carbolfuchsin (Primary stain) b. Acid-alcohol (Decolorizer) c. Methylene Blue (Counterstain)
Gram staining Gram (+) Gram (-)
Color Blue Pink Pink Red
Peptidoglycan Thick Layer Thin layer
Techoic acid in cell wall
Present Absent
Lipopolysaccharide in cell wall
Absent Present