1. Urine Testing

2. Urine testing... basement (basal) membrane acts as a filter on substances that have a relative molar mass > 69,000 This means that small substances 1.pass into the glomerular filtrate 2.if not reabsorbed will pass into the urine Urine can be tested for a number of substances: 1.human chorionic gonadotrophin (hCG) - pregnancy hormone 2.drugs such as anabolic steroids

3. Pregnancy testing

4. The implanted human embryo produces human chorionic gonadotrophin (hCG): 1.helps maintain production of progesterone by the ovary and then placenta (after 12 or so weeks) so…. 2.prevents menstruation The hCG hormone is a small molecule o relative molar mass of 36,000 o Fits through basal/ basement membrane o passes into the glomerular filtrate Its presence can be detected in urine using an over-the- counter monoclonal antibody dip-stick test

5. How do test strips work? The test strip contains monoclonal antibodies that are specific for: 1.hCG alone 2.the hCG-antibody complex 3.unbound hCG-antibody Key information: 1.The hCG-specific antibody is tagged with a dye (usually blue) and is mobile 2.The antibody against the hCG-antibody complex and against the unbound hCG are immobilised in separate sites at one end of the strip

6. Any hCG present in the urine binds to the hCG-specific antibody (usually blue) – forms a hCG-antibody complex – the complex moves along the test strip until it is trapped by a band of immobilised antibodies against the hCG-antibody complex – a (blue) line forms – indicates pregnancy. Unbound hCG-specific antibody also moves up the strip – it is also trapped by immobilised antibodies – antibodies against unbound hCG-specific antibodies are in a different position – So the blue line here indicates that the test is working properly – it should be present whether pregnant or not.. See diagram on next slide

7. 1 1. Test strip is dipped into a sample of urine or into a stream of urine. 2 2. Antibody moves up the test strip. 3 3. Blue stripe appears in the ‘control’ region showing that the strip is working but hCG is not present. Not pregnant. 4 4. Blue stripe appears in both the ‘control’ region and the test region showing hCG is present. Pregnant! Monoclonal antibodies against hCG are tagged with a blue bead. Control region with antibodies against unbound hCG-antibody. Test region with antibodies against the hCG-antibody complex.

8. Absorbent region with MAB against hCG (wee here) Control region shows the test is working. Test region: the result is not clear…. try again…

9. Anabolic steroids

10. Anabolic steroids... Anabolic steroids increase protein synthesis in cells  especially muscle cells  give a competitive advantage is some sports Anabolic steroids are banned in most sports because:  they give an unfair advantage  they have dangerous side effects e.g. liver cancer, heart disease and testicular atrophy Anabolic steroids are relatively small molecules  < 69,000  excreted in the urine They can remain detectible for a few days after their use has stopped

11. How do we test for them? Urine samples can be analysed for steroids using gas chromatography A sample is  vaporised and  passed with a solvent into the gas chromatography tube Different substances have different retention times in the chromatography tube  used to identify the substance