2.1 Microscopy Flashcards
When were light microscopes first developed?
16th-17th century
Describe cell theory (3 things)
- both plant and animal tissues are made from cells
- cells are the basic unit of all life
- cells only develop from existing cells
Cell theory timeline - Robert Hooke
1665 = Robert Hooke - thinly sliced cork, observed ‘cells’
Cell theory timeline - Anton van Leeuwenhoek
1674-83 = Anton van Leeuwenhoek developed powerful glass lenses, first to observe microorganisms and red blood cells, sperm cells and muscle fibres
Cell theory timeline - Dumortier
1832 = Dumortier observed cell division in plants, took years for theory to be accepted
Cell theory timeline = Robert Brown
1833 = Robert Brown first to observe plant nucleus
Cell theory timeline = Schleiden, Purkyne and Schwann
1837-38 = Schleiden proposed all plant tissues are made of cells
Purkyne was first to use a microtome to cut thin samples, proposed both animals and plants are made of cells
Schwann declared all living things are made of cells + their products (birth of cell theory)
Cell theory timeline - Remak
1844-55 = Remak observed cell division in animals, findings copied and published by Virchow in 1855 as his own
Cell theory timeline - Pasteur
1860 = Pasteur disproved theory of spontaneous generation of cells by showing that bacteria only grow in a sterile nutrient broth after being exposed to air
Why wasn’t cell theory fully developed before the mid-19th century?
Magnification of microscopes was not developed enough (high enough) to view cells in enough detail, theories not believed for many years
Name the two lenses on a compound light microscope
Objective lens and eyepiece lens
Describe the process of a dry mount
solid specimen, viewed whole or cut thinly, placed on slide with cover slip on top
Examples of dry mounting
E.g. pollen, hair, muscle tissue, plants
Describe the process of a wet mount
specimens suspended in liquid (water or oil), cover slip placed at an angle
Examples of a wet mount
E.g. aquatic samples, other living organisms
Process of squash slides
wet mount prepped first, lens tissue used to push down cover slip, good technique for soft samples
take care so cover slip isn’t broken
Examples of a squash mount
E.g. root tips - for viewing cell division
Process of a smear slide
edge of slide used to smear sample to create a thin, even coating on another slide
Examples of smear slides
E.g. blood samples - good to view cells
Why do scientists use stains?
Increases visibility , creates contrast between different organelles as they take up dye by differing amounts, can distinguish between different variations of organism e.g. bacteria
Crystal violet and methylene blue stains
Positively charged dyes which are attracted to negatively charged materials in cytoplasm which leads to staining of cell compounds
Nigrosin and Congo Red stains
negatively charged dyes and are repelled by negatively charged cytosol so stay outside of cells so they stand out against stained background
Differential staining
can distinguish between 2 organisms that are difficult to identify or can differentiate between organelles of a single organism
Gram stain technique - what does it do
used to separate bacteria into 2 groups - gram positive and gram negative
Gram stain technique - process
crystal violet dye added to specimen, then iodine which fixes the dye, washed with alcohol then a safranin dye is added to stain the negative bacteria which lost the blue stain
Gram stain technique - results if gram positive bacteria are present
Retain dye so appear blue/purple
Gram stain technique - results if gram negative bacteria are present
Thinner walls so lose the stain, then stained with a counter stain called safranin dye which makes them appear red
Gram positive bacteria + penicillin
Gram positive bacteria are susceptible to penicillin
Gram negative bacteria and penicillin
Gram negative bacteria have thinner cell walls so aren’t susceptible to penicillin
What is Acid-fast technique used to test for
used to differentiate species of Mycobacterium from other bacteria using a lipid solvent
Acid fast technique method
solvent used to carry carbolfuchsin dye into cells
cells are washed with a dilute acid-alcohol solution
then stained with a counter stain (methylene blue)
acid fast expected results
Mycobacterium aren’t affected by the acid-alcohol so retain the stain which is bright red
other bacteria lose the stain and are exposed to a methylene blue stain - turning them blue
Stages of slide prep - fixing
chemicals such as formldehyde are used to preserve specimens in a near-natural state
Stages of slide prep - sectioning
specimens are dehydrated with alcohols, placed in a mold with wax/resin to form a hard block which can then be sliced using a microtome knife
Stages of slide prep- why are specimens treated with multiple stains
specimens are often treated with multiple stains to show different structures
Stages of slide prep - mounting
specimens are secured to slide with a cover slip
Precautions around stains
Gloves, goggles, wash hands, clean surfaces, disinfect
Some stains are toxic or irritants so precautions must be taken
