Presentation on theme: "Measuring phenylalanine R Neil Dalton The WellChild Trust Laboratory King’s College, London Evelina Children’s Hospital at Guy’s Hospital NSPKU, Skipton,"— Presentation transcript:
Measuring phenylalanine R Neil Dalton The WellChild Trust Laboratory King’s College, London Evelina Children’s Hospital at Guy’s Hospital NSPKU, Skipton, 26 th February 2004
Measuring phenylalanine Phenylalanine The bad boy of phenylketonuria?
Measuring phenylalanine Phenylalanine is an essential amino acid One of the building blocks of all proteins Essential that the blood level of phenylalanine is maintained at an optimum concentration Optimum concentration range relatively wide, but in phenylalanine hydroxylase deficiency needs to be routinely monitored
Measuring phenylalanine phenylalanine 3D structure
Measuring phenylalanine phenylalanine structure
Measuring phenylalanine Collection of the blood spot is the most important step in obtaining an accurate measurement of phenylalanine in the blood How do we actually measure it?
Measuring phenylalanine product ion scan DETECTORDETECTOR Phe + Collision Cell MS2 IONS Q1 Q2 Q3 Q1 - selection of phenylalanine ion Q2 – fragment the phenylalanine ions Q3 – look for fragments of the phenylalanine
Measuring phenylalanine product ion scan
Measuring phenylalanine fragmentation
Measuring phenylalanine DETECTORDETECTOR Phe + Collision Cell x + IONS Q1 Q2 Q3 Q1 - selection of phenylalanine ion m/z 166 Q2 – fragmentation of phenylalanine ion Q3 - selection of specific fragment ion m/z 120
Measuring phenylalanine optimal phenylalanine control
Checking in l Samples posted in are received at the laboratory and are logged in to the computer.
Punching out spot l A special machine punches out the spots
Measuring phenylalanine Sample Preparation Punch 3mm diameter blood spot into deep-well (2ml) microtitre plate Add 150 l methanol/water (83:17) containing d5- phe and d4-tyr (5.6mmol/l) Seal plate with lid and mix for 20min at 300 cycles/min Place on Gilson autosampler at RT for injection
Mixing and shaking l Samples are then put onto plates and mixed.
Measuring phenylalanine poor phenylalanine control
results l Sample results are then checked by the laboratory staff.
Lab team l Our hard working biochemistry team.
Reported results l Our dietitians have both a phoned and faxed report of all the day’s phenylalanine samples received
Phoning parents l With all the results available, the dietitian phones the families, discusses both the child’s well being dietary changes if needed and often a bit of a social chit- chat.
Measuring phenylalanine A multi-disciplinary team of parents/individual, health care professionals, and leading edge analytical technology are involved in maintaining optimal blood phenylalanine Collection of the blood spot is the most important step in obtaining an accurate measurement of phenylalanine in the blood
Acknowledgements Charles Turner Sue Bird The WellChild Trust Charitable Foundation Guy’s & St Thomas’ Hospitals Guy’s and St Thomas’ NHS Trust