2. The placement
• Placement situated in Lyon, France
• A laboratory that deals with rare errors of
inborn metabolism
• Given the title “stagiaire” roughly translated
as “intern”.
• Duration of one year August 29th 2011-12
• Large cultural differences, language used was
French
3.
4. The organisation
• Service Maladies Héréditaires du Métabolisme
• Part of Hospices Civil de lyon
• Sectors on the floor comprised of biochemical,
radiological, enzymatic and biomolecular
laboratories
• Full time staff consists of a “cadre” of trained doctors
and management and also trained technicians.
• Public sector service with samples received from all
over mid-region France.
5. My responsibilities
• Performing technician duties such as:
– Carrying out the manipulations
– Responding rapidly to occasional emergency
samples
– Storage and cleaning of equipment
– Basic maintenance of machinery
– Removal of Hazardous waste
– Basic calculation and quantification
– Filing(paper) work
6. Classical organic acid disorders
• Manipulation consisted of dilution adjustment, urea removal,
adding of known concentration standards and silylation.
• Diagnosed by gas chromatography coupled with mass
spectrometry (enabling identification and quantification)
• Emergency cases in patients do occur (Hyperammonemia) so
some samples are prioritised for expendenicy.
• Detected by marker molecules that are the substrates (and
their toxic by-products) from a missing enzyme
• Examples of analytes included homogenisitic acid,
methylcitrate and methymalonic acid.
• Amino acid, carnitine and acyl-carnitine profiles commonly
ordered alongside.
8. Niemann-Pick C-1 Assay
• Cholesterol transportation failure in cells
• We made an attempt to functionalise a
biochemical assay for this disease from
an article
– Involved saponification of sample with KOH
(removal of FAs from cholesterol derivatives)
– A step of solid phase extraction in addition to
the normal liquid/liquid extraction
– Analysed by GC/MS
• Currently Niemann-Pick disorders (A,B,C
and D) are screened by cholesterol
studies on cultured fibroblasts using
immunocytofluorescence
10. Thin Layer chromatography
• Various Lysosomal storage disorders screened by TLC
• Oligosaccridosises (family of disorders)
• Sialic acid pathologies
• Adenylosuccinyl lyase deficiency
• Extremely old technique (60+ years)
• Enables identification and crude quantisation
• Consists of
– Ultra filtration of the sample
– Deposition of sample
– Migration of plate
– Revelation of bands
• Rf values are calculated from length of bands against a
control used
• Intensity of bands and depot judged by eye and
recorded
• Finally Rf values are compared with a reference table
to identify the bands.
11. Skills learned
• Laboratory conduct and expertise
• Practical professional knowledge of biochemistry
• Professional behaviour in a work-place
• French language learned
• Initiative when dealing with problems
• Improved team skills
• Perseverance
