8. Methods to study metabolism Flashcards
What is the principle of measuring metabolites?
Where you can measure and what (few examples): Blood - FFA, glucose, hormones Urine - urea, glucose, electrolytes, hormones Muscle - lactate, amino acids Liver - Glycogen, TGs, water Sweat - electrolytes fat Gut etc
What are the advantages and disadvantages to measuring metabolites
Pros
- Easy to sample
- Easy to measure
- Repeated sampling is possible
Cons
- Doesn’t give information on flux
- Does not give information about what happens in the tissues
What is the principle of Isotopic Tracer Methodology?
Add a known amount of tracer to an unknown tracee. Mix and take a small sample .
An isotope is stable if…
one or more extra neutron in the nucleus compared to the normal element
An isotope is radioactive if…
A neutron turns to a proton.
what are the pros and cons of using a stable isotope over a radioactive one
- safe, non-toxic
- Can be used in kids, pregnant women
- Many tracers can be used at the simultaneously in repeat studies
- Cost of some tracers very high
- Relatively large amounts of tracers needed
- Expensive equipment , technically qualified staff needed
what are the pros and cons of using a radioactive isotope over a stable one
- Ionising radiation
- Cannot be used in all populations
- Limitations in number of tracers and repeat studies
- No long lives radioisotopes for O and N
- No background problems, small amounts of tracer needed
- Cheaper and simple equipment
What is the principle of using AV-balance studies for metabolism?
= arterio-venous balance of stuff going into muscle and what comes out.
Mostly done in the leg as you have good access to the vein and artery and not much interruption from organs, like in the torso.
what are the pros and cons of using AV-balance
Pros
- Gives info on flux
Cons
- Invasive
- Difficult to measure blood flow accurately
- Error of flow rate can be 20%
- AV difference is often small and hard to detect.
What is the principle of using muscle biopsy as a method of studying metabolism?
Use a Bergstorm needle to obtain 20-100mg of muscle.
Numb skin. Numb fascia. Cut skin and fascia. Insert needle. Apply suction. Take Biopsy. Collect and analyse
what are the Pros and Cons of using muscle biopsy?
Pros
- Direct access to tissue of interest
Cons
- Invasive
- Limitations on number of samples
- Procedure may impact metabolism being sampled
What is NMR and the 2 forms of it?
Nuclear Magnetic Resonance
It includes MRS (Magnetic Resonance Spectroscopy) and MRI (Magnetic Resonance Imaging) - requires gradient coils.
What is the principle of MRS?
Non-invasive technique for tissue imaging. ATP, Lipid, Glycogen etc
Atoms with a single electron in their outer shell behave like small magnets. They line up with the direction of the magnetic field and with a radiofrequency pulse they can be made to take the opposite direction - when they fall back to their original state then emit a signal with a specific resonance freq- this is what is detected by the MRS
What are the pros and cons of MRS
Pros
- Non invasive
- Changes can be followed over time
- Several muscle groups can be looked at at the same time
Cons
- Expensive + uncommon
- Claustrophobia
- Experienced + skilled technicians needed
what is the principle of immunohistochemistry?
The process of localising proteins in cells of a tissue section.
It exploits the principle of antibodies binding specifically to antigens in biological tissues
- Primary antibody binds to specific antigen.
- Secondary antibody links to the primary
- Fluorescent probe is attached to the secondary antibody
