1.0.0 Development of practical skills in biology Flashcards
What is a light microscope?
- A type of microscope used to view cells by producing an enlarged image that shows some of the detail inside the cell.
- Uses different sets of lenses (compound microscope) to magnify the specimen.
- Does not include internal structures (limited detail)
What are light microscopes used for?
Include HOW
Examining microscopic organisms, cells or tissue sections using a light source.
Light source: visible light (EM)
Do not include the function
Different componenets of a microscope
There are 9 main components
- Arm
- Stage
- Eyepiece (ocular) lense
- Objective lenses
- Condenser
- Iris diaphragm
- Illuminator
- Coarse focus
- Fine focus
Functions of the microscope components
- Supports microscope parts, carries the microscope
- Slides can be held down by stage clips
- Used to look down the microspcope to view image (x10)
- Set of lenses of differing magnifications that rotate into place directly above the microscope slide (x4, x10, x40)
- Directs the beam of light from the source onto the specimen
- Alters the amount of light directed onto the specimen
- Light source used to pass light through the specimen
- Large focusing dial that is used to generally focus the image; used with low power lens only
- Small focusing dial that is used to focus the image sharply
Equation
Magnification
I
A M
Maginification = image size / actual size
Define resolution
The ability to see two distinct points seperately
What is the resolution of a light microscope
200 nm is the smallest distance at which two individua points can be seen as seperate objects
Conversions
1 nanometre (nm) = 10-9 m
1 micrometre (µm)= 10-6m
Produce a set of instructions on how to use a light microscope
- Place the low power objective lens in position (scanning/shortest lens x4).
- Place the microscope slide on the stage.
- Use the coarse focus to raise the stage close to the low power lens. It should lock into place.
- Use the coarse focus to slowly raise the low power lens until the object is visible and in focus.
- Use the fine focus to sharpen the focus on the image.
- Ensure that the part of the slide to be viewed is in the centre of the field of view.
- Rotate the medium power objective lens (x10) into place. It will be very close to the slide.
- Using the fine focus only, refocus the image. It should need very little adjustment but if needed go back to step 1.
- Repeat steps 7-8 with the high power lens (x40).
Ensure that
Always carry the microscope by the arm & support the base.
Decribe the setup of a cavity slide
A drop of pond water can be placed into the well in the slide to view live microscopic organisms such as protists (unicellular organisms) or Daphnia (small crustacean)
Describe the setup of a flat slide
Relates to thin structures
Used with thin sections (one cell thick) that are usually stained to enable structures to be seen and distinguished from each other.
Why is staining of tissues essential?
Cells & their components are transparent making it very difficult to distinguish them and their components as there is no contrast between them.
What is methylene blue stain?
Include what it does specifically
- It is used as a general biological
stain - It stains the nucleus blue
What is gram staining?
It is used to enable bacteria to be seen with the light microscope.
It stains gram positive bacteria violet and gram negative bacteria red
Detailed walkthrough of each step
Decribe the process of gram staining
What is H&E stain?
H&E stain is a mix of two dyes:
* Haematoxylin (blue)
* Eosin (red)
Decribe what each dye does
- Haematoxylin is a positively charged stain that reacts with negatively charged molecules in cells. (it stains acidic structurese blue)
e.g. it reacts with nucleic acids.
These stain blue - Eosin is a negatively charged stain and so reacts with positively charged molecules in cells. (it stains basic structures pink, i.e. cytoplasm)
e.g. it reacts with amino groups in proteins. These stain pink.
What is a stage micrometer?
A type of microscope slide with a scale etched onto it
* Used to calibrate an ‘eyepiece graticule’.
What is the eyepiece graticule?
Fixed scale inside the eyepiece lens. It must be calibrated for each objective lens using the stage micrometer.
Covert your answer into µm
What is one eyepiece division equivalent to (in mm) in this diagram?
One small division on stage micrometer = 0.1mm
One eyepiece division:
= 2 divisions on stage micrometer scale
= 0.2 mm
= 0.2 x 1000 = 200 µm
Different types of cells
Include examples
Relates to microscopy slides light micrographs
- Squamous cells e.g. wall of alveoli, lining of mouth
- Cuboidal cells e.g wall of kidney tubules
- Columnar cells e.g intestine lining
- Ciliated cells e.g lining of trachea
- Red and white blood cells e.g. lymphocytes
Name a type of microscope derived from a light microscope
Light microscopes can be used with different techniques to enhance contrast in the image - laser scanning confocal microscope
Magnification - around x10000
Resolution - approx. 1000nm
Decribe how it works?
Laser scanning confocal microscope
- A laser beam is passed through the objective lens to illuminate a single point in the specimen.
- Light photons released from the specimen are passed through a photomultiplier tube (PMT). This amplifies the light and converts it into a digital signal.
- Any unfocused light is filtered out by passing the light through a pinhole.
- The laser scans across the specimen point by point at a specific depth to build up an image pixel by pixel on the computer screen.
- Scanning at different depths enables a 3D image to be built up.
What is an electron microscope?
Type of microscope that uses a a beam of electrons to obtain an image.
