Microscopy Flashcards
Why is oil immersion useful?
- Clearer view of specimen
Due to light being refracted each time there is passage through medium
Light refracts each time it moves through any medium with varied refractive index (i.e. glass to air), this causes a reduction in image quality for every passage.
If we can minimize the number of passages then the image quality will be increased and resolving power preserved due to less refraction.
Immersion oil is formulated in such a way it that has identical refractive index to glass. Therefore no refraction of light occurs with movement from the glass to the oil.
Parfocal
When switching from lower to higher power lens, the image will stay in focus with minimal focusing necessary.
Why start with lowest objective lens?
When utilizing lowest magnification, the depth of focus is increased. So the lower the objective power, then high the depth of focus.
So when using high power, a smaller portion of specimen will be in focus.
As such it will be easier to locate and focus on an organism on low power then move to higher magnifiication.
Immersion oil should NEVER be used with the high dry lens i.e. 40x (T/F)
True
Once there is oil on the slide you CANNOT use the 40X lens— what is an alternative solution?
Rotate the nosepiece the other way to use 4X or 10X to refocus.
As magnification increases, the area of the field of view _________________, the depth
of the field of view _______________________, the working distance __________________, and the
amount of light required ____________________.
As magnification increases, the area of the field of view decreases, the depth of the field of view decreases, the working distance decreases, and the amount of light required increases.
How does magnification affect working distance?
The working distance decreases as you increase magnification.
The high power objective lens has to be much closer to the specimen than the low-power objective lens in order to focus.
How does magnification affect light intensity?
There is a fixed amount of light per area, and when you increase the magnification of an area, you look at a smaller area. So you see less light, and the image appears dimmer.
Morphology vs arrangement
While arrangement refers to the groupings of individual cells, morphology describes the appearance of groups of bacteria, or colonies
What would happen if we used 100x without immersion oil?
Light refracts each time it moves through any medium with varied refractive index (i.e. glass to air), this causes a reduction in image quality for every passage. Thus the total image quality will be significantly decreased due to refraction.
Common shapes of bacteria
Cocci, Spirals, Bacilli
Common arrangements of bacteria
- Cocci Coccu Diplococci Tetrad Sarcina (8) Streptococci Staphylococci - Spirals Spirochete Spirillum Vibrio - Bacilli Bacillus Diplobacilli Streptobacilli Palisades Coccobacillus
Can growth patterns be a definitive identification ?
Even on general purpose growth media, bacteria can exhibit characteristic patterns of growth.
While these growth patterns are an important piece of information when identifying a bacterial species, they are not sufficient for positive ID.
Staining procedures and metabolic tests must be used for a definitive identification
Simple stain -
Simple stain
- only one stain is used, and all types of bacteria appear as the color of that stain when viewed under the microscope.
E.g. crystal violet, safranin, and methylene blue. Simple stains can be used to determine a bacterial species’ morphology and arrangement.
Mycoplasma stains well with gram stain (T/F)
False; Mycoplasmas, which have no cell wall, stain poorly with the Gram stain
