Microscopy

Question 1

What are the four aspects of a microscope?

Answer 1

Detector, objective, specimen and a light source.

Question 2

Why are microenvironment created when using microscopes?

Answer 2

To ensure that the temperature, CO2 and humidity is maintained.
This prevents focus instability.

Question 3

How is this microenvironment maintained?

Answer 3

the box - custom design for individual microscopy. allows easy access
The cube - high quality fan, stops noise and vibration
Air tight table top - doesn’t allow any unwarranted air flow or entry of substances,, live cell cultures grow.

Question 4

Why should you consider time scales with microscopes?

Answer 4

different investigations take different time scales - microtubule takes seconds, developmental weeks.

Question 5

What is the triangle of frustration?

Answer 5

balance between temporal resolution, spatial resolution and sensitivity.
balance between the three gives the best picture.

Question 6

What 3 uses does light microscopy have?

Answer 6

Histology, phase contrast of fibroblasts cultures and time lapse microscopy.

Question 7

What are the two types of electron microscopy?

Answer 7

Transmission or scanning.

Question 8

How can fluorescence be used in microscopy?

Answer 8

fluorescence is created by molecules emitted light that they have absorbed. The absorbed light is what excites them

Question 9

What is stokes shift?

Answer 9

is the difference, in nanometres, between the peak excitation and the peak emission wavelengths. Each fluorophore has a distinct and individual Stokes Shift.

Question 10

What is photobleaching?

Answer 10

Attachment of specific fluorochromes to various parts of the cell to produce a coloured image of areas of interest.

Question 11

Name some common fluorochromes?

Answer 11

GFP, DAPI, FITC, TRITC and Cy5

Question 12

Name 6 uses of microscopy?

Answer 12

Immunohistology
Laser capture microdissection - removing the area of the cell that you want to focus on
Phase contrast- shows cell morphology
Time lapse - show cell migration
Tissue and cell localisation - shows live imaging of division and dispersal. (either intracellular live imaging - small scale cell migration and co localisation- visualising two components at once.
3D Z series add many images together to make a 3D shape reconstruction of a cell.