Microscopy And Fluorescence

Question 1

What is the stain Called that light microscopy uses

Answer 1

Gram stain

Question 2

What is gram stain dependant on

Answer 2

The outer membrane of gram negative bacteria

Question 3

What stain is used for TB testing

Answer 3

Acid fast stain

Question 4

Explain the steps of gram staining on light microscopy

Answer 4

1- both stained purple with crystal violet

2- iodine is used to set the violet into cell wall

3- short term decolorisation with alcohol causes gram -ve to become colourless

4- Safranin stain is added to turn gram -ve pink

Question 5

What instead of gram staining and light microscope is used to detect bacteria

Answer 5

PCR

Question 6

What is the biggest advantage of electron microscopy to light

Answer 6

They have a higher resolution due to electron shorter wavelengths

Question 7

Why do samples in electron microscopes have to be fixed and dehydrated

Answer 7

They are placed into a vacuum which kills cells

Question 8

What did the electron microscope help to find for the bacterial structure

Answer 8

It is split into 2 by the septum

Question 9

What are the issues with electron microscopy

Answer 9

Needs a thin sample to see high resolution

Forms artefacts

Not live - only dead cells used

Question 10

What is another word for Scanning electron microscopy to see 3D structures

Answer 10

Cryoelectron microscopy and tomography

Question 11

How are cells kept live in cryoelectron and tomography

Answer 11

They are frozen with ethene

Question 12

What is better about cryoelectron tomography than electron microscopy

Answer 12

Doesn’t need fixation or thin sectioning

Produced a 3D imagine

Question 13

What is an issue with cryoelectron tomography

Answer 13

Poor contrast

Question 14

What is the most recent microscopy technique used

Answer 14

Fluorescent microscopy and digital imagining

Question 15

What does fluorescent microscopy and digital imaging help do that no other does

Answer 15

It shows compartmentalisation in cell components through dyes

Question 16

What 2 ways are cell components dyed in fluorescent microscopy

Answer 16

GFP

Immunofluorescence

Question 17

What is the filter and mirror called which helps only blue light wavelength pass in fluorescent microscopy which helps visualise the dye

Answer 17

Emission filter

Dichroic mirror

Question 18

What is better in fluorescent microscopy which uses light compared to light microscopy

Answer 18

Much better resolution

Question 19

What is immunofluorescence and name 2 types

Answer 19

Detection through marked antibodies

Direct or indirect

Question 20

What is direct immunofluorescence

Answer 20

Monoclonal antibodies are attached to fluorescent molecules which bind to the target

Question 21

What is indirect immunofluorescence and why is it time efficient

Answer 21

Primary antibodies will attach with no fluorescent molecule

Secondary antibodies attach to them with a fluorescent dye which is easier to attach than to a primary antibody

Question 22

What is bad about immunofluorescence

Answer 22

Cells need to be fixed to allow antibodies to work (no live imaging)

Question 23

What is GFP tagging

Answer 23

Gene for a green fluorescent protein added onto gene of interest to track its expression over time

Question 24

How is GFP switched on

Answer 24

When the gene of interest is expressed in transcription mrna GFP also expressed

Question 25

What is good about GFP over immunofluorescence

Answer 25

It has live imaging

Question 26

What is an issue with GFP tagging

Answer 26

They can aggregate to other gfps on other genes

Question 27

Name 2 things which were found using the fluorescent microscopy technique

Answer 27

FTSZ - microtubules site for cell division

MreB- actin site when cell wall synthesis occurs