Apparatus and Techniques Flashcards
Colorimeter
An instrument used to quantitatively measure the intensity of a colour of a solution
Apparatus + method
- curettes are special rectangular test tubes - use a blank filled with distilled water to 0 each reading - make a calibration curve
Calibration curve definition
a method for determining the conc of an unknown solution by comparing it to a range of known concentrations
Calibration curve
- use serial dilutions of known concs of reducing sugar - heat with excess known conc of benedicts solution- use same volumes of solutions each time- zero colorimeter using a blank- read results using transmission/absorbance - plot on a graph = calibration curve- test unknown sugar in same way- colour change, blue -> green -> yellow -> red
Variables to control
- light intensity - wavelength of light - temperature - air movement - humidity
Explain results
as leaves are covered/ removed- surface area of leaves decreases - number of stomata decreases - transpiration decreases - cohesion-tension decreaseswhen all leaves covered or removed- some evap (not transpiration)- from upper surface of leaves
Sources of error
- water used to make cells turgid - water used by photosynthesis - water made by respiration - leaks in apparatus - not all stomata removed- stem not cut under water (allowing air into xylem which breaks cohesion-tension)- error in reading miniscus
Calculating rate of transpiration
(( distance bubble moved ) x ( cross sectional area of capillary tube )) / time taken
Serial dilution
progressively diluting a concentrated solution by known dilution
Ten fold serial dilution
- 1cm3 of initial- added to 9cm3 of distilled - mix until homogenous - 1cm3 transferred to next
How to use a light microscope
- Start with lowest power mag objective lens - put the slide onto a stage and hold in place using stage clips- use the course focus to move the stage as close to the lens as possible without touching - look through eye piece and move focus so stage moves away from the lens and comes into focus - adjust fine focus and light intensity for clearest image - repeat with higher power lens if necessary
What is the eyepiece graticule
see through ruler fitted inside the eyepiece
What is the stage micrometer
Slide with markings on it
How to work out actual size using a light microscope
1 - set mag2 - record how many eyepiece graticule units is equal to 1 micrometer 3 - don’t change mag4 - view specimen5 - use scale to work out actual size
micrometer to mm
1 micro = 0.001 mm
Slide prep
- Add a few drops of water to blank microscope slide - Use forceps to pick up sample- Thin enough sample so light shines through - Place onto microscope slide - Use pipette to add stain - Place cover slip on top - Put slide on microscope stage - Adjust mag
Staining
- Identify cellular components - Identify different cell types in a tissue - only see parts that absorb light - stains make more parts of the cell absorb light (become visible)- different stains for different cells
Nuclei stain
nile blue
Cytoplasm stain
easin
Cell membrane
sudan blue
General principle
- Sharp HB pencil- Clear continuous lines- No sketching- No shading- Draw only what you see- Square guidelines - Whole page - Rubber- Include titre - Include scale
Common mistakes
- Sketchy lines- Curved labels
Labelling
- Sharp pencil- Use a ruler - No arrow heads - No crossing lines- Annotate structures - Underline species name
Chromatography
Process used to separate mixtures of coloured complexes
Paper chromatography
Uses paper as the stationary phase
TLC
Uses a thin layer of absorbant methanol
How does chromatography work
- mixture contains more than one compound, each compound has a different solubility - the mobile phase moves through the stationary phase- the more soluble, the further it moves - compounds separate in order of solubility
Chromatography method
1 - Add conc spot of mixture to the bottom of stationary phase2 - End of chromatography strip is lowered into solvent 3 - Pigments/ compounds are carried different distances according to solubility4 - if compounds not visible use a stain5 - Stop before solvent reaches the end of the stationary phase6 - Identify compounds by comparing results to known sample (RF)
How to calculate RF
Distance travelled by compound / Distance travelled by solvent
Electrophoresis
Using electricity to separate DNA fragments by length
Process
- DNA negatively charged - attach a fluorescent label or stain DNA - put DNA fragments into well at negative end - turn on current - DNA fragments attracted to positive electrode - Small fragments move further faster- 3 fragments = 2 cuts
For
- very simple organisms - basic nervous systems
Against
- cannot consent - could be fatal - do not know what they feel- use of animals is wrong
Aseptic technique
Lab procedures that are performed under sterile conditions to prevent contamination
Importance of aseptic technique
- avoids contamination- no nutrient contamination - unchanged conditions - no yield decrease - prevent micro organism escape
Example of technique
1 - Disinfect work area2 - Flame inoculating loop after transfer 3 - Flame mouth of glass bottle 4 - Don’t put bottle tops on bench 5 - Quickly replace lid
Dissection safety
- Use scissors instead of scalpel where possible - Replace blade covers when not in use - Make cuts with blades facing away from you - Wear safety goggles- Sterilise equipment
What is needed for fast diffusion
- High surface area - Short diffusion distance - High concentration gradient
Adaptions for high surface area
- Branching e.g. trachea- Folding e.g. alveoli
Adaptions for short diffusion distance
- Thin membranes
Adaptions to maintain high concentration gradient
- Blood circulation - Ventilation of air
How to prevent water loss
- Waxy cuticle - Close trachea/stomata- Trap moist air around holes
How does trapping air around holes prevent water loss?
- Reduce water potential gradient - Reduce water vapour loss
