experimental biology Flashcards
Any additional context required
This Flash card serves only as a key, it is here only to help you identify which flash cards are for what practical.
Hints [Range of marks]
Practical name (strand 1 or 2)
Most of these flashcards are based off of the questions about the practicals in the past paper , not the practicals themselves.
Why is gas mixture bubbled through the krebs solution
Up to 3 reasons [1-3]
Rat/Guinea pig illeum (1)
- supply the cells with oxygen
- ensure rapid mixing of the organ bath
- keep the solution at the correct pH
If contstructing an appropriate concentration response curve for two neurotransmitters in the absence of an antagonist, what are the you required to have on the graph.
(drawing the graph will help you) [8]
Rat/Guinea pig illeum (1)
- correct title (in this case - Contraction of guinea pig uterine smooth muscle as a function of agonist concentration (acetylcholine or histamine) in absence of clemizole
- labelled x axis ( agonist/drug conc in log M)
- x axis values (** negative log M values in ascending order** )
- Labelled Y axis (Muscle contraction in g ( values start at 0)
- distinguishable between two neurotransmitters (diff colours/shapes)
- dots appropriately linked (smooth sigmoid curve)
- key (Neurotransmitter A & B w/o antagonist)
ANY graph question on this practical requires the EXACT SAME ANSWER
How would you decipher EC50 using a graph
[2]
Rat/Guinea pig illeum (1)
Look for the midpoint of the sigmoid curve/ when the Log M value is = to half of the maximal response
How would you decipher EC50 from a table
[1-2]
Rat/Guinea pig illeum (1)
by looking at log (M) value where the response value is half of the maximal response
(if maximal response is 7, look for where the response is 3.5)
If looking at a table, how would you be able to tell that the addition of an antagonist has had an affect on the EC50 value
[1-2]
Rat/Guinea pig illeum (1)
It would take longer to reach the maximal response/ the Log (m) value at which the response is half of the maximal has changed
ACh and histamine are the agonists. Their receptors are mAChR and H1
Assume the antagonist has had no affect on the EC50 of ACh but has affected the EC50 of histamine. The maximal response for both agonist actions remains the same.
What actions can be argued that the antagonist has taken on the receptors.
[3]
Rat/Guinea pig illeum (1)
The Antagonist does not antagonise the mAChR
but appears to be a competitive antagonist of histamine H1 receptors (1 mark),
because the Histamine EC50 with antagonist > EC50 without antagonist (1 mark)
How can you tell that a competitive antagonist has worked.
[1]
Rat/Guinea pig illeum (1)
It increases the EC50 of the agonist.
What is EC50.
General
The concentration required in order for an agonist of a receptor to ellicit half of its maximal response
Assume you wanted to test an antagonist on a New agonist and two seperate forms of treatment (A & B) were repeated 10 times each.
How would you statistically
determine whether the effects of the antagonist on the EC50 of the new agonist are significant or not? Which assumption are you making?
(this is in essence the full question) [4]
Rat/Guinea pig illeum (1)
- check whether data are normally distributed
- assuming they are , look at assumptions made based on how experiment was conducted. Should have two means of log EC50 therefore its a matter of paired or unpaired T-tests
- tests will be paired if experimenter used same illeum before and after - best set up as it removes natural variation
- test will be unpaired if we have 20 diff pieces of illeum
Why is glucose added to the krebs solution
[1]
Rat/Guinea pig illeum (1)
To supply the energy requirements of the tissue
You are provided with a 25 mM stock solution of acetylcholine. Explain how you would produce a final concentration of 40 nM (volume of 20ml) in the organ bath
CiVi=CfVf (Ci&Vi = conc and vol of stock solution, Cf&VF = dilute sol)[4]
Rat/Guinea pig illeum (1)
Vi=CfVf/Ci 40x10^-9 x 20x10^-3/25x10-3 = 32 x 10-9 l = 32 nl. (1 mark)
(the numbers are arbitrary, the point remains the same)
This is impossible to measure, (1 mark)
So need to make a dilution of 1/1000 so as to measure 32 μl of that new solution and add that volume in the organ bath (1 mark)
To make a 1:1000 dilution, add 1μl of 25mM stock into 999 μl of water (1 mark)
this exact solution doesnt matter, just know how to do this in general
Why is krebs solution used in this practical?
[2]
Rat/Guinea pig illeum (1)
- it has a similar ionic composition and osmolarity to tissue fluid
- it contains nutrients and energy to allow for cellular metabolism
The two agonists in this experiment are always histamine and ach
Describe how you might determine whether the effects of the antagonist on the EC50 for histamine are significant.
Rat/Guinea pig illeum (1)
Repeat the experiment with histamine ± clemizole (same concentration) 3 to 5 times [1 mark]
Get the means and standard deviation or standard errors of the mean of the EC50s without clemizole and of the EC50s with clemizole (2 marks)
and compare these two means via a 2 sample paired t-test (1 mark)
what are some reasons for taking multiple readings of DBP, PP & pulse prior to treatment
[3]
Pulses and Blood pressure (1)
- to account for normal variations within a subject’s pulse/ BP
- to account for nervousness/excitement of subjects at the start
- to account for students being inexperienced at taking measurements
- to allow subjects to recover between measurements (the sphygmomanometer cuff can be quite uncomfortable)
- that the spacing between measurements is the same as that after the subject has taken the drug, so these measurements are the control values.
dont need all of these, these are just the options
why is it important that the limb is relaxed when measuring pulse
[4]
Pulses and Blood pressure (1)
If muscles are contracted then muscle spindles twitch – this may be mistaken for the pulse (2 marks)
If a drug increases PP (pulse pressure) and decrease DBP (diastolic blood pressure) around the same amount, what can be identified
[2]
Pulses and Blood pressure (1)
PP = SBP - DBP
If the decrease in DBP is similar to the increase of PP that means that the SBP has not changed.
mention specific doses of when the drug is affecting these things
If plotting a graph on the effect of salbutamol on heart rate/pulse, What would the graph require
[7]
Pulses and Blood pressure (1)
- title - effect of salbutamol on heart rate
- x axis- time (min)
- y axis heart rate (bpm)
- dot to dot line joining (no line of best fit)
- can distinguish between before and after treatment time points of administration need to be indicated clearly with an arrow
- 95% confidence interval ranges if the question asks for it
Given that salbutamol is described as a beta-agonist, suggest why subjects often experience tachycardia after the repeated administration of salbutamol?
[3]
Pulses and Blood pressure (1)
Direct action on β1 -adrenoceptors on the cardiac pacemaker cells (SA node). [1 mark]
Direct action on β2-adrenoceptors on smooth muscle cells of blood vessels causes vasodilatation, reduced total peripheral resistance and therefore reduced diastolic blood pressure. [1 mark]
fall elicits a baroreceptor reflex (carotid sinus afferent limb; deceased vagus output and increased sympathetic output to the heart) that increases heart rate indirectly. [1 mark]
describe step by step how you would analyse the effect of giving a single dose of salbutamol to 6 subjects whos pulse was measured before and after administration. Determine whether salbutamol had a significant effect & what statistical test would you use
(basically a full question) [4]
Pulses and Blood pressure (1)
- analysis that considers the before and after measurements of each subject separately (natural pulse variations) [1 mark]
- For each subject, we need to calculate the ‘difference’ between values before and after treatment [1 mark]
- Calculate mean and standard error of the mean for the ‘differences’ values. [1 mark]
- hypothesise that if there is no effect, the difference should be equal to zero. [1 mark]
- Carry out a paired t-test to see if the mean of the ‘differences’ varies significantly from zero. Accept also carry out a one sample t-test to compare the mean of the differences to 0 (our null hypothesis value). [1 mark].
bacteria extracted from 10g samples ,100ul spread onto LB agar plates
why is 100ul of the final extract spread onto the agar plates
The wording of this is weird [2]
E coli (2)
We start from 100 g of meat that gets extracted into 1000 μl of LB (100 g / ml), so plating 100 μl (1/10) of the initial culture is equivalent to plate the bacteria present on 10 g of meat.
does it matter whether you prepare 200ul or 1000ul of your 10 fold dillution
[4]
E coli (2)
- no
- if doing it w/ 20ul, 20ul of final extract is added to 180ul of LB. Plating 100ul is equvalent to plate bacteria found on 1g of meat , just needs to be multiplied by 10 to get results for 10g
- if 100ul of original stock is used, add 900 ul of LB, plating 100ul is equivalent to plate bacteria found on 1g, just needs to be multiplied by 10 to get results for 10g
For the levels of how much bacteria is growing = 6 classifications, the lowest classification has 0 colonies, the largest has >8000
Explain the process of preparing various dillutions to discriminate the 6 classes of contamination. Assume the assay allows you to count reliably in between 2- 200 colonies per plate & that each class should be covered by at least two different dillutions
Also mention the tools you would require to do this (e.g P20, P100, P1000 pippettes)
| [8]
E coli (2)
- serial dillution with 1/10 incriments
- sample with a titre of 10,000 will be read accurately at dilutions 1/100 (100) and 1/1000 (10)
- w/ A P100 & P1000 can measure 20-100 & 200-1000 ul (& our max final vol is 1.5 ml)
- 1 (neat), 1/10 (100 μl of neat into 900 μl of LB), 1/100 (100 μl of 1/10 into 900 μl of LB), 1/1000 (100 μl of 1/100 into 900 μl of LB). or 1/5, 1/50 1/5000, 1/5000
there are 6 classifications in the amount of bacteria colonies
Distributor A
145 / 234 / 138
Distributor B
22 / 10 / 8
Warehouse A
5 / 0 / 8
Warehouse B
338 / 164 / 182
Restaurant A
12 / 8 / 16
Restaurant B
1804 / 756 / 2876
deduce which distributor is delivering ground meat to which warehouse and which warehouse is delivering meat to which restaurant.
full question, its not hard, just need to know [8]
E coli (2)
- little diff between Distributer B, Warehouse A and Restaurant A (1 mark)
*They all fall into the same class II of borderline contamination (1 mark). - Distributor A shows higher titre of class III level. (1 mark),
- Warehouse B is again higher, one class up in IV (1 mark)
- and the Restaurant B is the highest but still in class IV. (1 mark).
- It is most likely that Dist A delivers to Warehouse B which delivers to Rest B as there are all class II (1 mark)
- It is thus most likely that Dist B delivers to Warehouse A which delivers to Rest (1 mark), but it is unclear why the level of contamination would increase down the food supply chain. (1 mark)
there is obviously not all the info required to give this answer here. The main point is, you should be able to identify what the pathways would be & that they want you to do it in detail (list classifications etc)
what statistical test would you do to compare whether there is a significant difference between two sample means
[1-2]
General
two sample unpaired t test
How do you calculate mean cell volume
[2]
Haematology (1)
haematocrit (%) / red blood cell count (/L) = mean cell volume (fl)
how do you calculate mean cell haemoglobin
[2]
Haematology (1)
haemoglobin count (g/dL) / haematocrit{PCV} (%) = mean cell haemoglobin (g/L)
how do you calculate mean cell haemoglobin concentration (to 1dp)
[2]
Haematology (1)
haemoglobin count (g/dL) / red blood cell count (/L) = mean cell haemoglobin count (pg)
what is Haematocrit/Packed cell volume
[1/2]
Haematology (1)
Volume of red blood cells that are expressed as a proportion of the total volume of a blood sample.
Either expressed as a decimal or percentage.
What is the normal range for packed cell volume for females?
[1]
Haematology (1)
37-47%.
What is the normal range for packed cell volume for males?
Haematology (1)
40-54%
What is the cut off level for donating blood for men and women
[2]
Haematology (1)
125g of hb/L for women
135g of hb/L for men
what is mean cell volume , what are its units and what is its normal range
[2]
Haematology (1)
The mean volume of a single red cell.
femtolitres (normal range is 78-100fL).
What is the mean cell haemoglobin concentration, what is it measured in and what are its units
[2]
Haematology (1)
The mean quantity of haemoglobin per red cell.
The results are in picograms (pg), 1 pg is 10-12g.
The normal range is 27-32 pg.
What is the mean cell haemoglobin , what is it measured in and what are its units
[2]
Haematology (1)
The mean concentration of haemoglobin within the red cells (g/L).
The results are in g/L.
Normal range is 300-350 g/L.
If someone is in the blood group A, what antibodies would they have and what sera would you use to test this.
[1-2]
Haematology (1)
Anti-B antibodies
Anti-A sera (would see agglutination)
If someone is in blood group AB, what antibodies would they have and what sera would use to test this
[1-2]
Haematology (1)
No antibodies
Anti-A & Anti-B sera (would see agglutination in both)
If someone is in blood group O+, what antibodies would they have, what antigens would they have and what sera would use to test this
[1-2]
A & B antibodies
Rheuses D antgigens
Only agglutinate in anti-d sera
How would you plot a graph to show the proportions of people in the uk with each blood group?
[4]
Haematology (1)
You would do a histogram, a stacked column or a pie chat. labelled X and y Axis, & legend
How do you convert 0.85 sec into msec (as an example)?
ECG
Multiply by 1000 (850 secs)
When calculating heart rate using the equation 60/R-R intervals (secs), how would you round the answer?
ECG
Always round down because its only the observable beats per minute e.g., if the answer was 70.58, only 70 beats per minute were observed
What are R-R intervals?
ECG
The time between two consecutive R waves (peak of the QRS complex)
What is the relationship between the R-R and heart rate directly after exercise?
ECG
The R-R intervals will decrease in duration and the heart rate will increase (shorter R waves means more heart beats)
What is the correct order of the following events which are recorded by an ECG?
(1) Repolarisation of ventricles
(2) Depolarisation spreads across atria
(3) Sino-atrial pacemaker fires
(4) Depolarisation spreads through ventricular walls
ECG
3241
What is the P-R interval?
ECG
It is the time taken from the onset of the P wave to the start of the QRS complex.
(Time between atrial depolarisation and ventricular depolarisation).
What will happen post exercise to the P-R interval?
ECG
It will quickly go back as the heart rate slows.
Which circulatory systems do the left and right ventricles pump blood through?
ECG
Left:
- systemic circulation
Right:
- pulmonary circulation
How do action potentials fire in the heart?
ECG
ECG
In a wave, they are not fired simultaneously.
Name the three leads used in ECG and what parts of the body they are associated to.
ECG
Lead I:
- RA-LA
Lead II:
- RA-LL
Lead III:
- LA-LL
What is the T wave in ECG?
ECG
This is the repolarisation of the ventricles.
What are some abnormalities that ECG can help to diagnose?
Cardiac rate (tachy or bradycardias)
Arrythmias (rhythm problems)
Conduction in the heart (heart block/arrythmias)
What is meant by the latency period in the EMG practical?
EMG
The period between stimulation of the ulnar nerver at the elbow and the contraction of the little finger.
How do you calculate conduction velocity?
EMG
distance between the two sites and the time taken for the signal to reach the destination.
How do you calculate the conduction velocity?
EMG
you are given a distance, then find the time associated with it (if you are given the distance and the time just divide them to get the velocity).
If you are not given the distance for the exact time then you find the difference in times which will give you you length and the time taken.
How do you calculate the amplitude and the latency period before contraction of the finger on a graph?
EMG
Amplitude:
Measure from baseline and find the peak (difference is the amplitude)
Latency period:
Measure from the inception of the stimulus and the peak of the amplitude (difference in times on the graph is the period).
What are two characteristics of a nerve fibre that will increase its conduction velocity?
EMG
Increase the diameter and add myelination
Why is the stimulating electrode bathed in saline prior to applying electrical stimulation?
EMG
To improve conduction between the electrode and the skin.
How do you calculate making a solution of a certain % w/v?
EMG
w/v means g/100ml
so take the total required volume and find what volume would be the % by multiplying the total volume by the percent (e.g., 100ml x 0.09 (9%) = 9g/100ml)
(another example:)
750 of 0.9% w/v
750ml = 0.9 x 7.5 (100mls) = 6.75g
Why do motor units from different muscles contain differeing numbers of motor fibres?
EMG
Those with fewer motor fibres per neuron are concerned with more precise movements.
What produces striations of muscle?
EMG
Produced by regularly spaced myofibrils, which consist primarily of actin and myosin filaments.
The movement of these filament is responsible for contraction and relaxation of the muscle fibre.
How are muscles electrically connected to the spinal cord?
EMG
Via motor neurons which synapse with individual muscle fibres at a region called the neuromuscular junction.
What is the diameter and range in length in skeletal muscle fibres?
EMG
diameter of 100um
length range from a few mm to 30cm
What are the FVC, FEV1 and PEFR?
Lung mechanics
FVC: forced vital capacity
FEV1: Forced expiratory volume in 1 second
PEFR: Peak expiratory flow rate
When recording forced vital capacity, what is it essential for the volunteer to do?
Lung mechanics
Wears a nose clip.
Describe how to use a peak flow meter.
lung mechanics
- Ensure the scale and the holes in the end of the peak flow meter are not obstructed
- Move the scale marker back to zero.
- Fit a clean mouthpiece
- In either standing or seated position, the volunteer must breath in as deeply as possible and close their lips tighly around the mouthpiece.
- The volunteer must breather out as hard and as fast as possible.
Why must a participant to be standing or sitting when measuring the peak expiratory flow rate?
lung mechanics
This test requires the volunteer to breathe out fully and forcefully. Either sitting or standing removes any restriction on the ability of the diaphragm or external intercostal mucles to contract.
Therefore allows for the most accurate reading.
Why are three recordings are taken respectively for FVC, FEV1 and PEFR? Why is the higest value taken?
lung mechanics
The practice effect, volunteers become more familiar with the equipment and how to perfom the procedure the more they do it. Using the highest value takes into account that the volunteer might of been nervous or excited at the start of the experiment
What piece of equipement is used to measure FVC and what is the FEV1 calculated from?
lung mechanics
Force vital capacity is measured using a spirometer and the FEV1 is calculated from the FVC recording.
Know how to calculate mean and standard deviation and plot them onto graphs.
All practicals
this has come up on multiple practicals so go look if you are not sure.
What is PEFR - what does it show?
lung mechanics
Peak Expiratory Flow Rate - it measure the rate at which you breath air out.
What are some potential factors impacting the difference in lung mechanic indices recorded between volunteers?
Lung mechanics
Male/female
Height
athleticism
Pre-existing respiratory problems
Do they smoke/how long for?
What are the sampling rate and scale amplitude for recording vital capacity?
Lung mechanics
200/s sampling rate
100mV for scale amplitude
What can FEV/FEV1 tell us?
Lung mechanics
It is the volume of air forcibly expelled by the lungs in 1 second.
It is abnormally low in patients with obstructive diseases
70-75% is healthy average
How do you convert from molar concentrations (1 x 10^x) to pM, nM, uM and mM?
What are each of these units expressed in standard form?
Radioligand binding
1 x 10^-12 = 1pM
1 x 10^-9 = 1nM
1 x 10^-6 = 1uM
1 x 10^-3 = mM
Starting with a stock of 10mM, describe how to make up the stock concentrations of 1 x 10^-6 to 1 x 10^-11
Radioligand binding
Starting with stock solution 1 x 10^-2, make 100ul of each as only 2ul is needed per stock solution (will specify needed amount somewhere in the question).
To do this, take 10ul of starting stock 1 x 10^-2, and add 90ul of saline solution (or what is specified in question) - creating stock 1 x 10^-3
Repeat this with the new solution until you have the required stock concentrations.
What is IC50?
Radio ligand binding
It is how much antagonist/drug is needed to inhibit the response by half.
How do you calculate specific binding?
Radioligand binding
By subtracting the non-specific binding( predicted by the line on the drawn graph) from the total binding measured at each concentration of radioligand.
Expressed as a %
How to calculate the IC50 from a graph on a log scale?
Radioligand binding
Find the maximum response, go to half this point of the y axis. Read off the corresponding value on the x axis and put that as the power of 10 (10^x) this will give you the value.
Then convert to the required units.
In a radioligand experiment results graph, how can affinity be determined from the sigmoidal curves?
radioligand binding
The closer the curve is to the Y axis (the smaller the concentration (M) it binds at) the higher the affinity of the ligand.
What is the purpose of a radioligand binding assay?
Radioligand binding
Determines the binding affinity between endogenous liquids and their molecular targets using specific radiolabelled ligands.
What is the rate of association dependent on?
Radioligand binding
the concentration of the drug and the concentration of the receptor.
What are the equations for the law of mass action?
Radioligand binding
Rate of association: k1 = [D] x [R]
Rate of dissociation: k-1 = [DR]
What makes the system at equilibrium?
Radioligand binding
When the association rate = the dissociation rate (the number of molecules that are binding to the receptor is the same as the number of molecules that are dissociating from the receptor.
What is KD?
radioligand binding
The equilibrium dissociation constant
k-1/k1 = (D) (R)/ (DR) = KD (subscript D)
