Bright Field Microscope
An instrument that magnifies an object by passing visible light directly through the lenses and object.
Fluorescence Microscopy
An optical system on the light microscope that uses ultraviolet light to excite dye containing
objects to fluoresce.
Phase Contrast Microscopy
An optical system on the light microscope that uses a special condenser and objective lenses to examine cell structure.
Dark Field Microscopy
An optical system on the light microscope that scatters light such that the specimen appears
white on a black background.
Simple Microscope
1 Lens
Compound Microscope
Commonly used in lab setting
2 Lens - Ocular Lens (Closest to Eyes) 10X
Objective Lens (Closest to Specimen):
10X (Low)
40X (High)
20 X
100X (Oil)
Condenser
Group of lenses below the stage; functions as a light-gathering system which sends light to
the specimen from the light source
Iris Diaphragm
Iris-like closure system below the stage which regulates the amount of light passing through
the condenser
Resolving Power
The ability to distinguish two points as distinct, separate objects rather than as one
blurred image. Under oil immersion (1000X) this distance between points should be 0.22 um. Resolving power depends on the wavelength of light (shorter=better), the desi
Monocular Microscope
single ocular
Binocular Microscope
two oculars
Numerical Aperture
Relates to the size of the cone of light entering the objective and the medium
surrounding the objective (usually air)
Immersion Oil
Prevents loss of light rays due to diffraction because the oil had the same refractive
index as glass
Total Magnification
Ocular magnification multiplied by the objective magnification
(Example: 10X * 100X = 1000X total magnification)
Ocular lense
eyepiece - close to the eye
Objective lense
close to the object on the stage
Parfocalism
Once the specimen has been focused under low power (10X), the microscope is parfocal if
you are able to switch to higher magnification with a minimum of focal adjustment
Working Distance
The distance between the bottom of the objective lens and the slide. As the
magnification increases, the working distance decreases
Light Intensity
Less light is required at low magnifications. As the magnification increases, the need for
light increases. However, too much light can "burn out" the image
Diameter of the Field
As the magnification increases, the diameter of the field decreases
Real Image
Image passing into the objective lens from the specimen
Low-Medium Objectives
Spirogyra
Volvox
Rhizopus
Oil Immersion Objective
Sacchromyces
Mixed Bacteria
Virtual Image
Real image is further magnified by the ocular lens and passes to the retina of the eye.
Virtual image is upside down and reversed right to left
Coarse Adjustment Knob
Used to bring the specimen into approximate focus
Fine Adjustment Knob
Used to bring the specimen into clear focus. When the fine adjustment knob is
in mid-range, a white band is visible
Refraction
the bending of light as it passes from one medium to another - through the objective lens from the specimen to produce a real image
Resolution
clarity of an image
Scientific method
-Observation
-Background Research
-Construct Hypothesis
-Experiment and Collect Data
-Analyze the Results
-Draw Conclusions
Observation
-1st step
-quantitative (measurable) & qualitative (immeasurable)
-prefer quantitative: less subjective & likely to involve human error
Background Research
research the topic to ensure that you do not repeat experiments that have already been successful
Construct Hypothesis
Hypothesis or Null hypothesis; both must be testable & only one will be correct
Experiment
Design must follow these three rules:
-You must be able to repeat the experiment with similar results (Precise & Accurate)
-Only one variable should be tested at a time (Independent & Dependent)
-Must always include an experimental control (Negative & Pos
Collect Data
-As the experiment progresses you must collect date which will be analyzed.
-The data can be collected before, during, and after the experiment.
-The data can be quantitative and qualitative.
Analyze the Results
Determine if the experiment supports or disproves the hypothesis
This part of the scientific method can include:
-Calculations
-Conversions (time, etc.)
-Graphs
-Tables
Draw Conclusions
Based on your analysis of results you make a decision on whether to accept the hypothesis or reject the hypothesis.
Hypothesis
a statement describing what the scientist thinks will happen in the experiment
Null Hypothesis
testable statement that if proven true will prove the hypothesis incorrect
Independent Variable
these are the variables in the experiment that can be changed (amount of water, time of day)
Dependent Variable
these are the variables in an experiment that will change as a result to a change in the independent variable (color, rate of the reaction)
Negative Controls
samples you expect NO change in during an experiment (sterile swab used to inoculate a sterile nutrient agar plate)
Positive Controls
samples you expect change to occur during an experiment (swab with E. coli is streaked across a nutrient agar plate)
Significant Digits
Rules:
-Any non-zero number (1-9) is ALWAYS significant
-Anytime a zero appears between significant numbers, the zero is significant
-Zeros that are ending numbers after a decimal point or zeros that are after significant numbers before a decimal point ar
Scientific Notation
This method is used when dealing with very small or large numbers.
-Example
26000000 = 2.6 X 107
0.0000014 = 1.4 X 10-6
Percent of Error
used to analyze data when a correct value is known.
Percent Error = I(experimental - actual)I * 100%
actual
Lab Report
Once the scientific method is complete most scientists will compile their results into a lab report.
Components:
-Title
-Abstract
-Introduction
-Material & methods
-Results
-Discussion
-Conclusion
-References
Conversions
Temp: C=5/9 (F-32)
Time: 60s=1min; 60m=1hr
Mass: 1000mg=1g; 1000g=1kg
Length: 2.54 cm - 1 in
Accuracy
simply a measurement of how close a group of values/results are to the goal value/result; relative measurement
Precision
simply another way of describing repeatability or reproducibility; only thing that matters is that multiple attempts create the same result
Media
Purpose: allow scientists to grow microorganisms in the lab setting
Must contain:
-Carbon - plant or animal source
-Nitrogen - from a plant or animal source
-Phosphorus
-Sulfur
-Minerals
-Water
Types: Complex, Defined, Selective
Forms: Liquid or Solid
Complex (Undefined) Media
media where the exact makeup and amounts of components in the media are unknown
Defined Media
media where each component and its amount is known and controlled to facilitate the growth of specific organisms
Selective Media
media used to select for particular types of bacterial growth
Procedures for Sterilization
Autoclave & Membrane Filtration
Autoclave
An autoclave sterilizes using high pressure as well steam to kill contaminants
Membrane Filtration
This technique uses positive pressure as well as a filter to remove contaminants of certain sizes
Aseptic Technique
Technique used to transfer microorganisms from one inoculation plate to another WITHOUT contamination, or to add bacterial samples to media plates
Phase Plate
special filter; mounted in a phase objective lens & retards rays