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| Question | Answer |
| Concentration vs activity | Common features: both allow comparisons to be drawn between different drug samples |
| Concentration vs activity: Concentration | - Units: Moles - Preferred measure - Relies on knowing the molecular mass of the substance - Methods: Physiochemical eg mass spectrometry Immunoassay eg radioimmunoassay Bioassay eg 2x2 bioassay |
| Concentration vs activity: activity | - Used when concentration cannot be found- when molecular mass is not known with any certainty - "units of activity" can express the amount of protein present and be measured using a bioassay |
| Finding the concentration of a drug/hormone using a radio-immunoassay | 1. treat specialised dish coated in antibody with a urine sample containing unknown amounts of hormone 2. Wash 3. Treat with unknown amount of radio-labelled hormone 4. Wash 5. Count radioactivity *The amount of radioactivity present at the end of the process is inversely proportional to the amount of hormone present in the urine sample. AKA the more radioactivity measured, the less hormone in the sample* 6. A standard curve can then be constructed by repeating the experiment with known amounts of hormone. This curve shows the relationshipbetween radioactivity and hormone present, and from this concentration of hormone can be calculated. |
| Finding concentration of a drug/hormone using enzyme-linked immunosorbent assays (ELIZAs) | 1. Incubate sample in dish with antibody raised against hormone/drug to be measured. 2. Wash 3. Incubate with enzyme-linked secondary antibody raised against a different part of the hormone (against antigen for non-competitive, antibody for competitive). 4. Wash 5. Incubate with colourless enzyme substrate converted to coloured product |
| Key points of ELIZAs | - Relies on antibody/antigen interactions - Utilised 2 antibodies: primary and secondary - Measures colour change, so safer than radioactivity - Can detect antigen or antibody - Standard curves can be constructed for colour change in order to measure conc: 'amount' of colour is proportional to amount of enzyme-linked secondary antibody and the time colourless substrate in incubated for (set amount of time) - Most carried out in '96 well plates' |
| Bioassays | - A bioassay determines the activity/concentration of a drug in a sample by comparing response produced by the sample to a standard preparation: one of which is in Denmark, the other in London |
| Bioassays: 2+2 bioassay | - Simplest bioassay for determining the relative potency for 2 drugs. - If drugs are acting by the same mechanism, the middle portion of the concentration-response curves should be parallel. - 'M'=EC50test/EC50standard OR Log'M'=Log(EC50test)-Log(EC50standard) |
| Using a 2+2 bioassay to determine the concentration of drug in a sample | 1. Treat bioassay with a small (s1) & large (s2) volume of standard preparation 2. Calculate ratio of s1 and s2 (r) 3. Determine amount of 'test' producing 20% of max response (UI) 4. Add higher amount of unknown (UIxr) and check that it produces 20% of maximum response aka lines are parallel |
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