Chapter 2- 2.1- Microscopy Flashcards
Explain how a sample is prepared when the Dry Mount method is used?
Solid specimens are viewed whole, for example- hair, pollen, dust and insect parts,
Or cut into very thin slices with a sharp blade (called sectioning). For example- muscle tissue or plants.
The specimen is placed in the centre of the slide and a cover slip is placed over the sample.
Explain how a sample is prepared when the Wet Mount method is used?
Specimens are suspended in a liquid for example water or immersion oil.
A cover slip is placed on from an angle.
Aquatic samples and other living organisms can be viewed this way.
Explain how a sample is prepared when the Squash slides method is used?
A wet mount is first prepared.
Then a lens tissue is used to gently press down the cover slip.
Depending on the material, potential damage to a cover slip can be avoided by squashing the sample between two microscope slides.
This method is good for soft samples.
Care needs to be taken that the cover slip is not broken when being pressed.
For example, root tip squashes are used to look at cell division.
Explain how a sample is prepared when the Smear slides method is used?
The edge of the slide is used to smear the sample, creating a very thin, even coating on another slide.
A cover slip is then placed over the sample.
An example of a smear slide is a sample of blood. This is a good way to view the cells in the blood.
What do stains increase ?
Contrast as different components of the cell take up stains to different degrees. The increase in contrast allows components to become visible so they can be identified.
What are 2 examples of positively charged dyes?
Crystal violet
Methylene blue
Because they are positively charged dyes they are attracted to the negatively charged materials in the cytoplasm leading to the staining of the cell components.
What are 2 examples of negatively charged dyes?
Nigrosin
Congo
Because they are negatively charged dyes they are repelled by the negatively charged cytosol. Therefore these dyes stay outside the cells leaving the cells unstained, which then stand out against the stained background.
How is a sample prepared for staining?
Placed on a slide and allowed to air dry.
This is then heat fixed by passing through a flame.
The specimen will adhere to the microscope slide and will then take up stains.
What can differential staining distinguish between ?
Differential staining can distinguish between two types of organism e.g. bacteria that would otherwise be hard to identify.
It can also differentiate between different organelles of a single organism within a tissue sample.
What is Gram stain technique used for?
To seperate bacteria into two groups, Gram-positive and Gram-negative bacteria.
Explain the procedure in distinguishing between gram-positive and gram-negative bacteria.
What are the results of the procedure and how is the gram-positive bacteria different to the gram-negative bacteria?
What relevance is this In medicine today?
Crystal violet is first applied to a bacterial specimen on a slide.
Then iodine is added which fixes the dye.
The slide is then washed with alcohol.
The gram-positive bacteria retain the crystal violet stain and will appear blue or purple under a microscope. An example of this is Streptococcus pneumonia which causes pneumonia.
Gram-negative bacteria have thinner cell walls and therefore lose their stain. They are then stained with safranin dye, called a counterstain. These bacteria will then appear red or pink under a microscope. An example of this is Yersinia pestis which causes bubonic plague infecting humans and animals.
Gram-positive bacteria are susceptible to the antibiotic penicillin, which inhibits the formation of cell walls. Gram-negative bacteria have much thinner cell walls that are not susceptible to penicillin and therefore wouldn’t be stopped by this.
What is the acid-fast technique used to differentiate between?
How is the process carried out?
Species of Mycobacterium from other bacteria.
A lipid solvent is used to carry carbolfuchsin dye into the cells being studied.
The cells are then washed with a dilute acid-alcohol solution. Mycobacterium are not affected by the acid-alcohol and retain the carbolfuchsin stain (bright red).
Other bacteria lose their stain and are exposed to a methylene blue stain, which is blue.
How does a light microscope work?
A compound light microscope has two lenses- the objective lens, which is placed near to the specimen, and an eyepiece lens, through which the specimen is viewed. The objective lens produces a magnified image, which is magnified again by the eyepiece lens.
This objective/eyepiece lens configuration allows for much higher magnification and reduced chromatic aberration than that in a simple light microscope.
Illumination is usually provided by a light underneath the sample.
Opaque specimens can be illuminated from above with some microscopes.
