Chemical Pathology Services External User Guide
Contents
1. a iduronidase CV CCV CAC iduronate sulphatase CV CCV AF CAC heparan sulphamidase CV CCV CAC a glucosaminidase CV CCV CAC N acetyltransferase CV CCV CAC N ac galactosamine 6 sulphatase CV CCV CAC p galactosidase CV CCV CAC arylsulphatase B CV CCV CAC B glucuronidase CV CCV AF CAC multiple lysosomal hydrolases CCV AF CAC multiple lysosomal hydrolases CCV AF CAC multiple sulphatases CV CCV AF CAC Lipidoses GM2 gangliosidoses B galactosidase CV CCV CAC hexosaminidase A CV CCV CAC total B hexosaminidase CV CCV AF CAC galactocerebrosidase CV CCV CAC arylsulphatase A CV CCV CAC a fucosidase CV CCV CAC p mannosidase CV CCV CAC a mannosidase CV CCV CAC a N acetyl galactosaminidase CV CCV CAC neuraminidase CV CCV CAC a neuraminidase f galactosidase CV CCV CAC External Handbook This is a copy unless printed on controlled yellow paper Doc number CCL 003 Page 17 of 26 No unauthorized amendments or photocopies to be made Version number 1 5 Department of Chemical Pathology Great Ormond Street Hospital for Children a galactosidase CV CCV AF CAC Other lysosomal disorders External Handbook Other disorders B glucosidase CV CCV CAC sphingomyelinase CV CCV CAC acid esterase CV CCV CAC B si2. Medium or saline Do not freeze 5 10 ml li hep whole blood Send whole blood at 4 6 4 6w w ambient temp to reach lab within 18 h of collection muscle liver biopsy Freeze immediately 4 6 w Send frozen skin biopsy into culture Send at ambient temp 4 6 w Medium or saline Do not freeze muscle biopsy Freeze immediately 4 6 w Send frozen 5 10 ml lip hep whole Send whole blood at 4 6 w blood ambient temp to reach lab ideally within 24 h of collection liver biopsy Freeze immediately 4 6 w Send frozen muscle biopsy Freeze immediately 4 6 w Send frozen 5 ml li hep whole blood Send whole blood at 4 6 w ambient temp to reach lab ideally within 24 h of collection liver biopsy Freeze immediately 4 6 w Send frozen 5 10 ml lip hep whole Send whole blood at 4 6 w blood ambient temp to reach lab ideally within 24 h of collection liver biopsy Freeze immediately 4 6 w Send frozen Pompe CRIM testing contact the Enzyme Laboratory Tel 020 7405 9200 ext 2509 2440 Glycolytic enzymes 5 ml li hep whole blood Send whole blood at ambient temp to reach lab ideally within 24 h of collection muscle liver biopsy Freeze immediately 4 6 w Send frozen FATTY ACID OXIDATION DEFECT HYPOGLYCAEMIA External Handbook This is a copy unless printed on controlled yellow paper Doc number CCL 003 Page 11
3. Creatine kinase isoenzymes ve Dill CRIM testing ES Cystine ss LS CYSUNOSIS 13 D Disaccharidase Deficiencies esseseeeeeeee 19 DNA Analysis Dysmorphic screen Fu c sido a 13 G Galactocerebrosidase eese esee 13 Galactokin se nice ne e HORE EISE ERE EUER 10 Galactose 1 phosphate uridyl transferase 10 Galactose 1 Phosphate 10 Galactosialidosis iiti etes 13 Galactosiddse niei iE EE Reden 13 Gangliosidoses 13 Gaucher 13 Globotriaosylceramide 12 Glucose 6 phosphate hydrolase esse 10 Glucose 6 phosphate translocase ees 10 Glucosidase cniin pet Ha HO ciii ote 13 Gluc ronidase ii iiie oe berti 13 Glycerol kinase 19 Glycogen brancher 1l Glycogen debrancher 11 Glycosaminoglycans eerte 12 H Heparan sulphamidase eene 13 Hepato splenomegaly screen eee 15 He xosamuiridase irri ertet er tete ane aiia 13 HUMEN 13 p n 13 FIV 20 Hydroxy butyrate BOHB 12 Hypophosphalasia eet rere eripere 19 I Teel rretan ine On OR 13 Iduronate s
4. Sachs B1 variant hexosaminidase A Sandhoff total B hexosaminidase Niemann Pick A sphingomyelinase Sialidosis a neuraminidase Galactosialidosis a neuraminidase B galactosidase Krabbe leucodystrophy galactocerebrosidase Angiokeratoma screen For patients with an angiokeratoma Fabry a galactosidase Fucosidosis a fucosidase Sialidosis a neuraminidase Galactosialidosis a neuraminidase p galactosidase Adult Gy gangliosidosis p galactosidase a Mannosidosis a mannosidase p Mannosidosis B mannosidase Schindler a N acetyl galactosaminidase Aspartylglucosaminuria aspartylglucosaminidase DNA Analysis External Handbook This is a copy unless printed on controlled yellow paper Doc number CCL 003 Page 15 of 26 No unauthorized amendments or photocopies to be made Version number 1 5 Department of Chemical Pathology Great Ormond Street Hospital for Children The Enzyme Laboratory works closely with the Clinical Molecular Genetics Laboratory at Great Ormond Street Hospital to offer mutational analysis for many of the lysosomal storage disorders It is essential to test for the presence of the polyA mutation encoding a pseudodeficiency of arylsulphatase A in all patients with low arylsulphatase A activity For other disorders the Enzyme Laboratory will advise if mutational analysis is available
5. and or appropriate when a diagnosis is made External Handbook This is a copy unless printed on controlled yellow paper Doc number CCL 003 Page 16 of 26 No unauthorized amendments or photocopies to be made Version number 1 5 Department of Chemical Pathology Great Ormond Street Hospital for Children Prenatal diagnosis is available for the following disorders It is important that the diagnosis in the index case has been confirmed in an appropriate tissue The tissues suitable for assay are stated in the table It is essential to contact the Enzyme Laboratory 020 7405 9200 ext 2509 before taking any samples for prenatal diagnosis to discuss your requirements and transport arrangements For chorionic villus specimens it is our policy to assay the villi directly where appropriate and then to check equivocal results or confirm diagnosis of an unaffected fetus on cultured cells Direct and cultured cell assays are charged separately and an additional charge is made for the cell culture For amniotic fluid samples where the assay is performed on cultured cells the cost of the cell culture is charged additionally Lysosomal storage disorders Mucopolysaccharidoses mucolipidoses and multiple sulphatidosis Following amniocentesis electrophoresis of amniotic fluid glycosaminoglycans GAGs is carried out on all pregnancies at risk for a mucopolysaccharidosis mucolipidoses II and lll or a multiple sulphatidosis
6. are available to GOSH Clinicians provided by External Referral Laboratories details of which can be found in the Internal User s Guide CCLO02 Sample reception Chemical Pathology Reception Level 1 Paediatric Laboratory Medicine Camelia Botnar Building Great Ormond Street Hospital for Children Great Ormond Street London WC1N 3JH Prof Simon Heales Professor of Clinical Chemistry 020 7813 8321 DD simon heales gosh nhs uk Laboratory Director Clinical Lead Enzyme Metabolic Unit Director of Newborn Screening Ms Helen Aitkenhead Principal Clinical Scientist 020 7813 8318 DD helen aitkenhead gosh nhs uk Deputy Laboratory Director Deputy Clinical Lead Bleep 020 7405 9200 Blood Sciences Special Routine QA POCT bleep 0200 Mr David Wells Pathology Lead Laboratory Manager 020 7813 8320 DD david wells qosh nhs uk Mrs Katie Harvey Principal Clinical Scientist 020 7405 9200 x 7843 n e Bainbridge Head of Newborn Screening katie harve osh nhs uk Enzyme Laboratory Bleep 020 7405 9200 bleep 0397 Ms Helen Prunty Principal Clinical Scientist 020 7813 8319 DD helen prunty gosh nhs uk Metabolic Laboratory Bleep 020 7405 9200 bleep 2257 Dr Steve Krywawych Principal Clinical Scientist T 020 7405 9200 x 6758 steve krywawych gosh nhs uk Mr Daley Aolofaju Chief Biomedical Scientist 020 7405 9200 x 0214 daley aofolaju gosh nhs uk Blood Sciences Special Routine External Handbook This is a copy unless printed on con
7. assays 2 Perchlorate precipitation INDEX External Handbook This is a copy unless printed on controlled yellow paper RWW 20 20 20 21 22 24 25 Doc number CCL 003 Page 2 of 26 No unauthorized amendments or photocopies to be made Version number 1 5 Department of Chemical Pathology Great Ormond Street Hospital for Children Chemical Pathology Laboratory Great Ormond Street Hospital for Children NHS Foundation Trust GOSH is a CPA accredited laboratory providing a wide range of Chemical Pathology analyses with a special interest in the diagnosis and monitoring of inborn errors of metabolism The laboratory is fully staffed between 9 am and 5 30 pm Monday to Friday and staff will be available for any enquiries you may have For sample requirements and general enquiries not dealt with by this guide or for results please contact the helpline in the first instance For other enquiries advice on investigations clinical advice and interpretation or to request an urgent analysis the duty biochemist is available on bleep 020 7405 9200 hospital switchboard bleep 0589 The on duty clinical staff member can also be contacted by a long range message pager via the hospital switchboard out of hours For additional information see website http www labs gosh nhs uk This handbook contains of all the tests currently performed in house by the Department There are a number of investigations that
8. frozen LYSOSOMAL STORAGE DISORDERS LSD Initial investigations monitorin Glycosaminoglycans U 5 ml fresh random urine Send at ambient temp by up to 4 w special post Sialic acid U 5 ml fresh random urine Send at ambient temp by up to4w special post Ceramide trihexoside U 5 ml fresh random urine Send at ambient temp by up to4 6w CTH special post Globotriaosylceramide GL3 GB3 Tetrasaccharides U 5 ml fresh random urine Send at ambient temp by up to4 6w hex4 special post Tetraglucose glc4 Vacuolated B 2 ml EDTA whole blood Send at ambient temp by Contact lymphocytes see page 22 special post done in histopathology Histopathology Individual enzyme Assays available individually for the diagnosis of lysosomal storage disorders are listed below with samples suitable for the assay Turnaround is 4 6 weeks External Handbook This is a copy unless printed on controlled yellow paper Doc number CCL 003 Page 12 of 26 No unauthorized amendments or photocopies to be made Version number 1 5 Department of Chemical Pathology Great Ormond Street Hospital for Children Disease Assay Tissue Mucopolysaccharidoses I Hurler a iduronidase WBC F Il Hunter iduronate sulphatase WBC P F II A Sanfilippo A heparan sulphamidase WBC F IIIB Sanfilippo B a N acetyl glucosaminidase WBC P F II C Sanfilippo C N acetyltransfer
9. sampling collection UDP galactose RBC 2 ml li hep whole blood Send whole blood at 4 6 w No transfusion prior 6 ambient temp to reach w Contact lab prior lab ideally within 24 h of sampling collection Galactose 1 RBC 2 ml li hep whole blood Send whole blood at 4 6 w Phosphate ambient temp to reach lab ideally within 24 h of collection Fructose 1 P aldolase L liver biopsy Freeze immediately Send 4 6 w frozen without thawing Glycogen Storage Disorders GSD la Glucose 6 L Fresh liver biopsy Contact enzyme lab prior 4 6 w phosphate hydrolase to sampling Do not freeze Ib Glucose 6 E Fresh liver biopsy Contact enzyme lab prior 4 6 w phosphate translocase to sampling Do not freeze I a 1 4 glucosidase BS Blood spots 5 ml li Send whole blood at 4 6 w External Handbook This is a copy unless printed on controlled yellow paper Doc number CCL 003 Page 10 of 26 No unauthorized amendments or photocopies to be made Version number 1 5 Department of Chemical Pathology Great Ormond Street Hospital for Children hep blood EDTA blood for vacuolated lymphocytes ambient temp to reach lab ideally within 24 h of collection 5 10 ml li hep whole blood Send whole blood at 4 6 w ambient temp to reach lab ideally within 24 h of collection muscle liver biopsy Freeze immediately 4 6 w Send frozen skin biopsy into culture Send at ambient temp 4 6 w
10. use a plastic bottle or bag Ice should not come into direct contact with the specimen container to avoid risk of contamination or labels becoming illegible Place the ice and specimen s in a plastic outer container and seal with waterproof tape Include sufficient ice to cover any possible delays in delivery Ice packs are suitable for a journey time of less than 6 hours However DO NOT place ice packs from 20 C freezer immediately next to whole blood or cells Dry Ice Solid CO Pack the specimens as above The outer pack must be an insulator e g expanded polystyrene State CONTAINS SOLID CO on the outside Seal outer case with tape Include sufficient solid CO to cover any possible delays in delivery External Handbook This is a copy unless printed on controlled yellow paper Doc number CCL 003 Page 5 of 26 No unauthorized amendments or photocopies to be made Version number 1 5 Department of Chemical Pathology Great Ormond Street Hospital for Children First class post When sending specimens by first class post the packaging MUST comply with UN3733 packaging regulations and postal regulations The package must be labeled PATHOLOGICAL SPECIMEN and may only be sent 1 class letter post Where first class post is indicated this assumes that delivery will be made by the next day Please DO NOT POST on Friday or before a UK Bank Holiday Courier or express delivery A reliable service should b
11. yellow paper Doc number CCL 003 Page 26 of 26 No unauthorized amendments or photocopies to be made Version number 1 5 Department of Chemical Pathology Great Ormond Street Hospital for Children
12. Hospital for Children CARBOHYDRATE METABOLISM DISORDERS Congenital Disorders of Glycosylation CDG Initial investigation serum transferrins isoelectric focusing at Institute of Neurology If type pattern CDG 1a amp 1b should be excluded first If patterns are abnormal but clinically indicated then measure enzymes Phosphomannomutase WBC 5 10 ml well mixed li hep Send whole blood at up to 6w whole blood with no clots ambient temp to reach lab ideally within 24 h of collection F skin biopsy into culture Send at ambient temp up to 10w medium or saline by courier Do not freeze Phosphomannose WBC 5 10 ml well mixed li hep Send whole blood at up to 6w isomerase Whole blood with no ambient temp to reach clots lab ideally within 24 h of collection F skin biopsy into culture Send at ambient temp up to 10w Medium or saline by courier Do not freeze Galactose Fructose metabolism Disorders Reducing substances 5 ml fresh random Freeze immediately Sugar urine Send frozen chromatography Galactose 1 RBC 2 ml li hep whole Send whole blood at 2 4w phosphate uridyl blood ambient temp to reach transferase No transfusion prior 6 lab ideally within 48 h of Gal 1 PUT W collection Galactokinase RBC 2 ml li hep whole blood Send whole blood at 4 6 w No transfusion prior ambient temp to reach 6w Contact lab prior lab within 24 h of
13. INHS Great Ormond Street Hospital for Children NHS Foundation Trust Chemical Pathology Services External User Guide Updated August 2012 New website www labs gosh nhs uk CPA Sa Accredited Medical Laboratory External This is a copy unless printed on controlled yellow paper Doc number CCL 003 Handbook Page 1 of 26 No unauthorized amendments or photocopies to be made Version number 1 5 Department of Chemical Pathology Great Ormond Street Hospital for Children Reference No 0250 INTRODUCTION Senior staff Department sections and phone numbers REQUESTING SAMPLE COLLECTION Storage Packing Transport TURNAROUND TIMES NEWBORN SCREENING FACTORS affecting performance of biochemical investigations SUDDEN INFANT DEATH MORIBUND CHILD Biochemical investigations ASSAY DIRECTORY Amino acid disorders Carbohydrate Metabolism Disorders Fatty acid oxidation defect hypoglycaemia Lactate pyruvate disorders Lysosomal storage disorders Prenatal Diagnosis Organic acid disorders Peroxisomal disorders Urea cycle disorders Other inherited metabolic disorders Hypophosphatasia Disacharidase deficiencies Glycerol kinase deficiency Neuroblastoma screen Other tests Antimullerin hormone AMH Busulphan Inhibin B Trace elements copper zinc selenium manganese Vitamin A E Isoenzymes macroenzymes alkaline phosphatase amylase creatine kinase Renal tubular markers APPENDICES 1 Special enzyme
14. alic acid CV CCV AF CAC cystine CV CCV CAC a glucosidase CV CCV CAC Neuronal ceroid lipofuscinoses NCL1 CLN1 palmitoyl protein thioesterase CV CCV CAC tripeptidyl peptidase CCV CAC Glycogen storage disorders a glucosidase CV CCV CAC brancher CV CCV CAC Urea cycle disorders ornithine carbamoyl transferase fetal liver carbamoyl phosphate synthase fetal liver E arginase FB argininosuccinate lyase FB C release of CO from leucine CV This is a copy unless printed on controlled yellow paper Page 18 of 26 No unauthorized amendments or photocopies to be made Version number 1 5 Department of Chemical Pathology Great Ormond Street Hospital for Children Doc number CCL 003 ORGANIC ACID DISORDERS Organic acids incl 5 ml fresh random urine Freeze immediately Send frozen 2 4w methylmalonate N acetylaspartate 5 ml fresh random urine Freeze immediately Send frozen 2 4w 2 ml amniotic fluid By arrangement Send by courier as arranged Biotinidase 0 2 ml li hep plasma Freeze ASAP Send frozen 1 3w PEROXISOMAL DISORDERS Very long chain 0 5 ml li hep plasma Separate immediately Send by 1 2 4w fatty acids class post includes phytanate amp pristanate UREA CYCLE DISORDERS Amino acids P 0 5 mllihep Separate ASAP Freeze 1 2w plasma immediately Send froz
15. as WBC F IID Sanfilippo D N acetyl glucosamine 6 sulphatase WBC F IVA Morquio A N acetyl galactosamine 6 sulphatase WBC F IVB Morquio B p galactosidase WBC F Vl Maroteaux Lamy arylsulphatase B WBC F VII Sly p glucuronidase WBC P F Multiple enzyme defects Neuronal ceroid lipofuscinoses Batten disease Mucolipidosis Il I cell multiple hydrolases P VL F Mucolipidosis III pseudo Hurler multiple hydrolases P VL F Multiple sulphatidosis multiple sulphatases WBC P F Gangliosidoses Gy gangliosidosis p galactosidase WBC VL F Gm2 gangliosidoses Tay Sachs B1 variant hexosaminidase A WBC P F Sandhoff total B hexosaminidase WBC P F Leucodystrophies Krabbe galactocerebrosidase WBC F Metachromatic arylsulphatase A WBC F Glycoproteinoses Fucosidosis a fucosidase WBC P VL F a Mannosidosis o mannosidase WBC P VL F B Mannosidosis B mannosidase P WBC F VL Schindler a N acetyl galactosaminidase P WBC F Sialidosis a neuraminidase WBC VL F Aspartylglucosaminuria aspartylglucosaminidase pF Galactosialidosis a neuraminidase B galactosidase WBC VL F Other lipid storage disorders Fabry a galactosidase WBC P F Gaucher B glucosidase WBC F chitotriosidase P Niemann Pick A amp B sphingomyelinase WBC VL F Wolman amp cholesteryl ester storage acid esterase WBC VL F disease CESD Infantile INCL NCL1 CLN1 palmitoyl pr
16. dry ice freezing mixture Take the biopsy as many cores as possible minimum two and put it immediately in the boat it should freeze immediately and thereafter should be not allowed to thaw at any time Wrap up the core in the foil and put it in the pre chilled container making sure that the cap is tight and immediately replace in the deep freeze 40 C or lower A small part should be put into glutaraldehyde and if necessary some into formalin but the majority should be frozen for chemistry and enzymology Skin Biopsy See fibroblasts External Handbook This is a copy unless printed on controlled yellow paper Doc number CCL 003 Page 8 of 26 No unauthorized amendments or photocopies to be made Version number 1 5 Department of Chemical Pathology Great Ormond Street Hospital for Children Codes ASSAY DIRECTORY Tests highlighted in turquoise are analysed in the Metabolic Laboratory Test highlighted in green are analysed in the Enzyme Laboratory Tests highlighted in yellow are analysed in the Routine Laboratory AMINO ACID DISORDERS Plasma Urine CSF Blood spot Homocysteine Plasma Urine Succinylacetone Sulphite Sulphocystine branched chain 0 5 ml li hep plasma Separate ASAP Freeze immediately Send 1 2w frozen 5 ml fresh random urine Freeze immediately Send frozen 3 6w 0 2 ml clear CSF Freeze immediately Send frozen 1 2w 4 blood spot
17. e esses 19 Orotic acid ente tested reo intet torti 19 P Palmitoyl protein thioesterase eee 13 Phosphoethanolamine eee 19 Phosphofructokinase eese 11 Phosphoglucomutase 11 Phosphomannomutase 10 Phosphomannose isomerase 10 Phosphorylase 5 etr RE Ie ERR S HERR 11 Phytanate Pristanate Pyr Vale 5 eicere citri iege R Reducing Substances iren ttere reet tenebre tia eh ieu 10 Retinol binding Protein eeeeeeeeeneeene 21 S Sandhoff cured uera iota Arten iet E Sc 13 SanfillDDO c6 uet annee deed nete in decade 13 wma ELE 13 Selemulit octo simon m ere Re PEE 20 Sialic acid 12 13 18 ADDED 13 yp EET 13 Sphingomyelin se oe epo RH Eno eet 13 Succinylacetone Sugar chromatography esee 10 19 20 Sulphite 2 2 oraret dri er ente bent 9 Sulphocystine 5 o epe ee tee e ee E 9 T Tay SACHS eoo eite etait oth 13 Tripeptidyl peptidase I 13 U UDP palatose eedem ene eed editos 10 V Vacuolated lymphocytes eese 12 Very long chain fatty acids sse 19 Mirunum e EE E E 20 Vitamin E NVM A scene ee t a EE W Wolman amp cholesteryl ester storage disease CESD 13 Z VA HM 20 External Handbook This is a copy unless printed on controlled
18. e used and instructed to take the specimens to the Reception in Chemical Pathology in the Camelia Botnar Building Turnaround time given is the anticipated time taken between sample receipt and report under normal operating conditions Where the assays are batched and performed infrequently the time is given as a range up to the maximum anticipated time Time taken for sample transport and posting the report should be added to this Where appropriate abnormal results will be phoned or faxed to the sending laboratory In cases where results are required more urgently please contact the department to discuss your requirements prior to sending specimens so that samples can be fast tracked Blood spot assays to screen for phenylketonuria PKU congenital hypothyroidism medium chain acyl coA dehydrogenase deficiency sickle cell disorders and cystic fibrosis in the neonatal period are available Sample requirement 4 good blood spots collected between day 5 and 8 day of birth day 0 ona standard screening card dried at room temperature and enclosed in a glassine cover Please provide the dates of birth and sampling Send at room temperature by post immediately Results will be available within 4 working days External Handbook This is a copy unless printed on controlled yellow paper Doc number CCL 003 Page 6 of 26 No unauthorized amendments or photocopies to be made Version number 1 5 Department of Chemical Pat
19. en Organic acids U 5 ml fresh Freeze immediately Send frozen 2 4w includes orotic acid random urine N acetylglutamate Discuss with up to 6 w Synthase enzyme lab Arginase RBC 5 ml li hep Send whole blood at ambient up to 6 w whole blood temp to reach lab ideally within 24 h of collection L liver biopsy Freeze immediately Send frozen up to 6 w Argininosuccinate L liver biopsy Freeze immediately Send frozen up to6 w Synthase Argininosuccinate RBC 5 ml li hep Send whole blood at ambient up to 6 w Lyase whole blood temp to reach lab ideally within 24 h of collection L liver biopsy Freeze immediately Send frozen up to 6 w Carbamoyl L liver biopsy Freeze immediately Send frozen up to 6w Phosphate synthase Ornithine carbamoyl L liver biopsy Freeze immediately Send frozen up to6 w Transferase OTHER INHERITED METABOLIC DISORDERS Hypophosphatasia Phospho U 5 ml fresh Freeze immediately Send frozen 3 6w ethanolamine random urine Disaccharidase Deficiencies Enzymes jejunum 2 mg jejunum Snap freeze in liquid N2 Send up to 8 w biopsy frozen on solid dry ice Also see appendix 1 Sugar stool walnut size Freeze immediately Send frozen 3 6w chromatography stool Glycerol kinase deficiency Organic acids U 5 ml fresh Freeze immediately Send frozen 2 4w includes glycerol random urine Glycerol kinase F skin biopsy into Send at room temperature up to 10w culture medium Do not freeze or saline Ex
20. ent or 200 ul blood in EDTA This test is performed in the Dept of Histopathology at GOSH Tel 020 7829 8663 Fax 020 7813 1170 External Handbook This is a copy unless printed on controlled yellow paper Doc number CCL 003 Page 22 of 26 No unauthorized amendments or photocopies to be made Version number 1 5 Department of Chemical Pathology Great Ormond Street Hospital for Children U urine Send 5 ml urine Keep frozen until dispatched and send by 1 class post This is used for our metabolic assays not enzymes Dilute urines creatinine lt 1 0mmol L and infected urines pH gt 8 0 are unsuitable L liver M muscle J jejunal Contact the enzyme laboratory for instructions before taking liver and muscle biopsies as some assays require the biopsy in an unfrozen state These assays are only available with prior arrangement and when the tissue sample can be delivered to this laboratory within 1 hour after being taken Unfrozen samples must be transported in a sealed container on wet ice For most enzyme assays including disaccharidases a frozen biopsy is required After wrapping in aluminium foil the sample must be frozen immediately using solid CO or liquid nitrogen then placed in a labelled plastic bag The sample must be stored and transported frozen It is essential that the sample remains frozen at all times until it is assayed External Handbook This is a copy unless printed on c
21. g additional tests and sample retention If the sample is still available and sufficient in volume and is viable additional tests may be added by phoning the Helpdesk On occasion the requestor may be asked to send a further request form with details of the test required Samples are retained in accordance to the Guidelines published by the Royal College of Pathologists and the Institute of Biomedical Science The retention and storage of pathological records and specimens 4 edition 2009 All samples are stored for a minimum of 48 hours after the report has been issued most samples are stored for at least two weeks and many are stored for longer periods Please contact the Helpdesk for further advice The packing requirements for samples are specified under each analyte further on in the booklet General and room temperature All specimens must be in leakproof containers Seal cap of container with parafilm or similar waterproof tape Wrap each container with sufficient absorbent material to completely absorb the contents in case of breakage There should be no contact between containers Place the container s and packing in plastic bag and seal the bag Place the sealed bag together with the request form in a rigid fibre or plastic outer case The outer case should be sealed with tape NOTE the request form must not be inside the plastic bag with the specimen Ice Pack specimens as above Place the ice in a leak proof container
22. hology Great Ormond Street Hospital for Children What samples It is important to check the fluid in which the metabolites of interest most obviously accumulate e g urine for organic acids The next part of this booklet indicates the sample type required for the investigations offered When indicated e g because of metabolite instability it is necessary to make arrangements with the laboratory prior to collecting the sample When The time of the sample collection is crucial where characteristic metabolites accumulate only intermittently in the samples Whenever possible patients should be investigated during periods when they are unwell Samples should be taken as soon as possible after admission before changes in treatment and diet lead to the disappearance of relevant metabolites Sample integrity Bacterial activity in poorly preserved samples produces a rise in pH and can lead to both the appearance of bacterial metabolites and the breakdown of important components especially sugars and some amino acids Samples with a high pH may not be analyzed for this reason Faecal contamination of urine produces a similar effect Dilute urine makes the detection of urinary constituents unreliable and samples with creatinine concentration gt 1 mmol L are preferred Diet Some metabolic disorders are related to a particular dietary intake or are produced only in the fasting state Investigations should be carried out as far as possible on
23. idase p galactosidase Fucosidosis a fucosidase o Mannosidosis a mannosidase cell disease cell screen External Handbook This is a copy unless printed on controlled yellow paper Doc number CCL 003 Page 14 of 26 No unauthorized amendments or photocopies to be made Version number 1 5 Department of Chemical Pathology Great Ormond Street Hospital for Children p Mannosidosis p mannosidase MPS VII Sly p glucuronidase Multiple sulphatidosis arylsulphatase A Aspartylglucosaminuria aspartylglucosaminidase Schindler a N acetyl galactosaminidase Plasma chitotriosidase is assayed in all patients to exclude Gaucher disease Hepato splenomegaly screen For those patients with hepatomegaly and or splenomegaly suspected of having a lysosomal storage disorder Gy gangliosidosis p galactosidase Sialidosis a neuraminidase Galactosialidosis a neuraminidase B galactosidase Gaucher p glucosidase Niemann Pick A amp B sphingomyelinase Wolman amp CESD acid esterase Fucosidosis a fucosidase a Mannosidosis a mannosidase cell disease cell screen B Mannosidosis B mannosidase MPS VII Sly B glucuronidase In all patients with hepato splenomegaly plasma chitotriosidase is assayed Cherry red spot screen For patients with a cherry red spot on the macula Gy gangliosidosis p galactosidase Gm2 gangliosidoses Tay
24. l clinical details especially presence or absence of neurological features hepatosplenomegaly dysmorphic features are given on the request form so that appropriate assays can be carried out Please let us know if the mother is pregnant as we can advise on prenatal diagnosis Sample requirements Enzyme assays are classified under separate diagnostic groups with abbreviations for samples where appropriate These abbreviations are explained below WBC leucocytes for white cell enzymes Unless specified enzymes are assayed according to the clinical details given Blood transfusion within 4 weeks may interfere with the result and sampling at this time is best avoided if possible Send 5 10 ml well mixed blood in lithium heparin minimum of 5mls The sample must not contain any clots heparinise the syringe if the patient is difficult to bleed Send the whole blood sample to reach the laboratory ideally within 24 hours of sample collection the shorter the interval the better the quality of the sample For most enzymes up to 48 hours is acceptable However WBC cystine it is essential that the sample is received within 24 hours Send by courier or Royal Mail Special Next Day delivery to arrive before 14 30 on a normal working day Please avoid sending samples on a Friday in case of delays in transport The turnaround time for these assays is approximately 6 weeks RBC erythrocytes Blood transfusion in the previous 6 weeks invalidates resul
25. m Freeze immediately Send 3 6w chromatography urine frozen walnut size stool Vitamin A S 0 5 ml serum Protect from light 2 4w Separate serum ASAP Send by first post Vitamin E S 0 5 ml serum Separate serum ASAP 2 4w Send by first post Zinc P 0 4 ml li hep Separate plasma ASAP 1 2w plasma Send by first class post ISOENZYMES Alkaline phosphatase P S 0 5mlli hep plasma serum Send by 1st class post up to 4w isoenzymes Amylase isoenzymes P S O 5mlli hep plasma serum Send by 1st class post up to 5w Creatine kinase P S 0 5mlli hep plasma serum Separate and freeze up to 4 w isoenzymes plasma serum immediately after External Handbook This is a copy unless printed on controlled yellow paper Doc number CCL 003 Page 20 of 26 No unauthorized amendments or photocopies to be made Version number 1 5 Department of Chemical Pathology Great Ormond Street Hospital for Children collection Send frozen RENAL TUBULAR MARKERS External Handbook 1 ml fresh random urine Freeze soon after 1 3w collection 1 ml fresh random urine Send by 1 class post 1 3w This is a copy unless printed on controlled yellow paper Doc number CCL 003 Page 21 of 26 No unauthorized amendments or photocopies to be made Version number 1 5 Department of Chemical Pathology Great Ormond Street Hospital for Children It is important that ful
26. of 26 No unauthorized amendments or photocopies to be made Version number 1 5 Department of Chemical Pathology Great Ormond Street Hospital for Children standard card 3 B Hydroxy P 0 3 ml li hep plasma Freeze immediately Send 1 2w butyrate BOHB frozen Provide glucose result Free fatty acids P 0 3 ml li hep plasma Freeze immediately Send 1 2w non esterified frozen Provide glucose result fatty acids NEFA Acetoacetate B perchloric acid Freeze immediately see 1 3w supernatant appendix for protocol Send frozen Organic acids U 5 ml fresh random Freeze immediately Send 2 4w urine frozen Acylcarnitines BS 4 blood spot on Send by first class post 1 2w Diagnostic fast All the above investigations to be carried out at the beginning and end of the fast under close medical supervision in a Hospital unit experienced in carrying out these tests not advisable in patients under 18 months or under 5 kg in weight LACTATE PYRUVATE DISORDERS Lactate P 2 ml fluoride oxalate plasma Separate plasma assay 6h ASAP B perchloric acid precipitation Freeze immediately 1 2w see appendix for protocol Send frozen CSF 0 2 ml clear CSF Freeze immediately 1 2w Send frozen Pyruvate B perchloric acid precipitation Freeze immediately 1 2w see appendix for protocol Send frozen CSF perchloric acid precipitation Freeze immediately 1 2w see appendix for protocol Send
27. ontrolled yellow paper Doc number CCL 003 Page 23 of 26 No unauthorized amendments or photocopies to be made Version number 1 5 Department of Chemical Pathology Great Ormond Street Hospital for Children o For each sample prepare 2 tubes each with 500 ul of ice cold 0 46 mol L perchloric acid keep cold at the bedside on an ice pack Collect blood into a lithium heparin tube or CSF into a plain tube and IMMEDIATELY pipette 100 ul of the sample into each of the perchloric acid tubes Mix vigorously transport to the laboratory on the ice pack Centrifuge within 10 minutes at 4 C 3000 rpm for 5 minutes Freeze supernatant in separate tubes and transport frozen Any delay in sample precipitation will result in rapid deterioration of the analyte level Our method requires that the proportion and concentration of perchloric acid is strictly adhered to in order to produce reliable results Manufacturers supply perchloric acid at a variety of strengths Please prepare the working perchloric acid as specified below NB For B hydroxybutyrate acetoacetate ratio a separate unprecipitated plasma sample should be sent Stock perchloric acid Supplied by manufacturer 60 w w SG 1 54 70 w w SG 1 70 Keep the working reagent in a plastic bottle at 4 C External Handbook This is a copy unless printed on controlled yellow paper 2 50 ml stock made up to 50 ml with distilled water 1 94 ml stock made u
28. otein thioesterase WBC F Classic late infantile LINCL NCL2 tripeptidyl peptidase WBC F CLN2 Transport defects Cystinosis cystine WBC F Sialic acid storage sialic acid U VL F NB Prenatal diagnosis is available for these disorders External Handbook This is a copy unless printed on controlled yellow paper Doc number CCL 003 Page 13 of 26 No unauthorized amendments or photocopies to be made Version number 1 5 Department of Chemical Pathology Great Ormond Street Hospital for Children Grouped Enzyme Screens for Lysosomal Disorders The lysosomal storage disorders can be grouped according to clinical features and a group of enzyme assays can be carried out on a single blood sample which provides both white blood cells and plasma for analysis The clinical signs of a lysosomal storage disease may eventually develop to give a classic picture but diagnosis at an earlier stage can be more difficult e g while Type Il Gaucher disease leads to hepato splenomegaly neurological signs may be more obvious initially To meet this and other concerns all patients have plasma chitotriosidase measured to exclude Gaucher disease and other LSDs Palmitoyl protein thioesterase and tripeptidyl peptidase which are deficient in infantile INCL NCL1 CLN1 and classic late infantile LINCL NCL2 CLN2 neuronal ceroid lipofuscinosis are assayed in all patients under 16 years with neurological problems and al
29. p to 50 ml with distilled water Preparation of 0 46 mol L perchloric acid Doc number CCL 003 Page 24 of 26 No unauthorized amendments or photocopies to be made Version number 1 5 Department of Chemical Pathology Great Ormond Street Hospital for Children A PCOLOAC CALE m sucess 12 Acetyl galactosamine 6 sulphatase eee 13 Acetyl galactosaminidase sel3 Acetyl glucosamine 6 sulphatase Acetyl glucosaminidase oe Acetylglutamate Synthase esee Acetyltrabsferdse socie ceteri E EERENS Acid esteras e Acid MAltASE Em Acylcarnitines esses Alkaline phosphatase isoenzymes Amino acids neri ise Amylase iSOCNZYMES ccceccecceseeeseeseceeeeseeeeeesecseeeeeeeeneeeaees Antimullerian hormone eene 20 LNUNEDM 19 Argininosuccinate lyase seen 19 Argininosuccinate synthase e Arylsulphatase rtr ttn tn rrt trino oett eo cose 13 Aspartylglucosaminidase eee 13 Aspartylglucosaminuria eee 13 B Biotitud se iie pene ninth ine te Es 19 Busulphan 2 eig oe ee i nr ere eei ieee 20 C Carbamoyl phosphate synthase esses 19 Ceramide trihexoside CTH Cherry red spot screen n Chitotriosid se 4 doce etre t retorica COPPET
30. rror of metabolism is suspected in an infant who died suddenly collect the following samples as soon as practicable to minimize post mortem changes blood spots bile spots plasma urine CSF aqueous humor Blood stained CSF and urine should be spun and separated immediately and this should be recorded Freeze at 20 C Skin biopsy can also be taken see fibroblasts Please discuss the request with the duty biochemist 020 7405 9200 bleep 0589 before sending the sample The diagnosis of metabolic disease cannot be made after death unless the correct specimens have been appropriately collected If metabolic disease is suspected and the child seems likely to die before a diagnosis can be made it would be advisable to collect the following specimens Blood 10 ml in a heparinised tube Separate plasma promptly Freeze the bulk of the plasma the remaining plasma and red cells should be kept at 4 C Urine 20 ml in a plain container and deep freeze Blood spots for acylcarnitine Bile spots for acylcarnitine DNA f the condition is one in which DNA studies are likely to be helpful take 10ml blood into an EDTA tube and deep freeze the whole blood Tissue biopsies liver muscle heart Label the plain container with the type of tissues prior to taking the biopsies Pre cool a plain container in the deep freeze Obtain dry ice liquid nitrogen or a freezing pack Make a small boat with a piece of aluminium foil and place it on the
31. s on std card Send by first class post 4d 0 2 ml li hep plasma Separate ASAP Freeze immediately Send 1 2w frozen 5 ml fresh random urine Freeze immediately 3 6w 5 ml fresh random urine Freeze immediately Send frozen 2 4w 5 ml fresh random urine Freeze immediately Send frozen 72h 5 ml fresh random urine Freeze immediately Send frozen 3 6w Methionine load for the diagnosis of homocystinuria Preparation Pre load Methionine load Fast overnight 6 hours for infants At the beginning of the test empty bladder and discard sample Collect 1ml blood in a lithium heparin tube separate and precipitate immediately Empty bladder Give L methionine 100 mg kg body weight orally over 5 minutes in a flavoured drink or as tablets Collect 1 ml blood in lithium heparin tubes at 2 4 and 6 hours On each occasion separate and precipitate immediately Collect all urine passed over the 6 hours after giving the methionine load Send plasma supernatant and urine on solid CO for amino acid analysis Maple syrup urine disease enzyme diagnosis Leucine decarboxylating system Skin fibroblasts Send a skin biopsy sample in the culture medium See Appendix 1 External Handbook This is a copy unless printed on controlled yellow paper Doc number CCL 003 Page 9 of 26 No unauthorized amendments or photocopies to be made Version number 1 5 Department of Chemical Pathology Great Ormond Street
32. samples taken at the time the patient was symptomatic Dietary restrictions or feeding may cause characteristic metabolites to disappear and result in false negative results Dietary metabolites may interfere with organic acid amino acid or carbohydrate chromatograms Patients receiving intravenous amino acid mixture may have amino aciduria amino acidaemia or organic aciduria Information on the type of diet and the timing of the sample in relation to meals will aid in the interpretation of these complex analyses Drugs Drugs influence metabolic investigations by analytical interference or by modifying metabolic processes Details of all medication should be provided with metabolic investigations Exchange transfusions blood transfusions These may affect the analytes measured in blood and especially erythrocytes When requesting tests in such patients check whether adequate time has lapsed since the last transfusion For assays of enzymes and metabolites in erythrocytes the time interval should be 6 weeks Other factors include Age of specimen Time of specimen separation Specimen storage Specimen haemolysis icterus and lipaemia Fasting state External Handbook This is a copy unless printed on controlled yellow paper Doc number CCL 003 Page 7 of 26 No unauthorized amendments or photocopies to be made Version number 1 5 Department of Chemical Pathology Great Ormond Street Hospital for Children If an inborn e
33. so in adult patients if these disorders are suspected It is important that the laboratory is given full clinical details in order to carry out the appropriate combination of tests Turnaround time is 6 8 weeks Note Some diseases may present under more than one heading Neurodegenerative screen Evidence of neurological regression hypotonia fits etc Gy gangliosidosis p galactosidase Gy gangliosidoses Tay Sachs B1 variant hexosaminidase A Sandhoff total B hexosaminidase Krabbe leucodystrophy galactocerebrosidase Metachromatic leucodystrophy arylsulphatase A Fucosidosis a fucosidase a Mannosidosis a mannosidase p Mannosidosis p mannosidase Schindler a N acetyl galactosaminidase MPS VII Sly B glucuronidase cell disease cell screen Plasma chitotriosidase is assayed in all patients to exclude Gaucher disease All patients under 16 years of age are tested for Infantile neuronal ceroid palmitoyl protein thioesterase lipofuscinosis INCL NCL1 CLN1 Classic late infantile neuronal ceroid tripeptidyl peptidase lipofuscinosis LINCL NCL2 CLN2 Dysmorphic screen The first line test for a dysmorphic child is screening for a mucopolysaccharidosis by urine GAGs The following enzymes are indicated if a mucopolysaccharidosis is excluded Gm1 gangliosidosis p galactosidase Sialidosis a neuraminidase Galactosialidosis a neuramin
34. ternal Handbook This is a copy unless printed on controlled yellow paper Doc number CCL 003 Page 19 of 26 No unauthorized amendments or photocopies to be made Version number 1 5 Department of Chemical Pathology Great Ormond Street Hospital for Children NEUROBLASTOMA SCREEN HVA 5 ml fresh Freeze ASAP Send frozen random urine VMA 5 ml fresh Freeze ASAP Send frozen random urine OTHER TESTS Antimullerian P S 0 5 ml EDTA Separate and freeze plasma upto2m hormone AMH plasma or serum immediately after serum collection Send frozen Busulphan P 1 ml EDTA Arrange with lab prior to Same day blood at 0 0 5 sampling 1 1 5 2 4 7h Send sample on ice Must be pre immediately to local Lab booked Separate and freeze plasma ASAP Label samples clearly with time of collection Copper P 0 4 ml li hep Separate plasma ASAP 1 2w plasma U 10 ml aliquot of Collect 24h urine into acid 2 4w 24h washed container Note 24 h volume Inhibin B P S 0 5 ml li hep Separate and freeze plasma upto3m plasma or serum immediately after serum collection Send frozen Insulin S P 0 3 ml Separate plasma Send on ice 1d serum plasma Manganese P 0 5 ml whole Send whole blood by first class 2 4 w blood in Trace post metal container Selenium P 0 4 ml li hep Separate plasma ASAP 2 3w plasma Send by first class post Sugar U F 5 ml rando
35. trolled yellow paper Doc number CCL 003 Page 3 of 26 No unauthorized amendments or photocopies to be made Version number 1 5 Department of Chemical Pathology Great Ormond Street Hospital for Children Mr Derek Burke Chief Biomedical Scientist 020 7405 9200 x5290 derek burke gosh nhs uk Enzyme Laboratory Mr Ade Ifederu Chief Biomedical Scientist 020 7405 9200 x5290 ade ifederu gosh nhs uk Newborn Screening Ms Julie Leakey Chief Biomedical Scientist T 020 7405 9200 x5290 julie leake osh nhs uk Metabolic Laboratory Departmental Office results enquires 020 7405 9200 ext 5076 020 7829 8624 fax Email chempath pa nhs net Helpdesk general enquires 020 7405 9200 ext 5009 Metabolic Laboratory Enzyme Laboratory 020 7405 9200 ext 5225 020 7405 9200 ext 2509 2440 Email gos tr ENZYME nhs net Specimen Reception Routine Laboratory 020 7405 9200 ext 5009 Newborn Screening Laboratory 020 7829 8383 DD Email gos tr enquiresgosnbs nhs net A request giving the following information must accompany the specimen apart from newborn screening tests a minimum of three identifiers are required Patient ID Specimen Assay s required Clinical details Sender surname or family name forename or personal name date of birth many reference ranges are age dependent sex some reference ranges are sex related patients reference i e Hospital number laboratory NHS n
36. ts Send 2 ml heparinised blood to arrive in the laboratory within 24 hours of sample collection except for galactokinase epimerase which has to be assayed on the day of sample collection and should be arranged with the enzyme laboratory at least a day in advance Send by courier or Royal Mail Special Next Day delivery P plasma I cell screen etc Send 1 ml plasma from a lithium heparin blood sample to reach the laboratory within 24 hours of collection Send by courier or Royal Mail Special Next Day delivery F fibroblasts from skin biopsies Taking a skin biopsy Proceed under aseptic conditions Have sterile culture medium ready The forearm and axilla are suitable sites Swab the skin with alcohol or chlorhexidine not iodine or betadine Approximately 0 2 ml to 0 4 ml of 0 5 lignocaine or similar local anaesthetic is injected intradermally and just subcutaneously Take a 3 mm punch biopsy full thickness skin or ellipse 4 mm x 2 mm immediately transfer the skin to the culture medium IN EXCEPTIONAL circumstances sterile dextrose saline may be used Keep at 4 C or room temperature DO NOT FREEZE and send by courier or datapost Fill the container to the top to avoid any airlock Storage 4 8 C for 24 hours in sterile saline 3 to 5 days in sterile culture medium It will take up to 6 weeks to grow fibroblasts VL vacuolated lymphocytes Send 3 unstained and 1 stained blood film made and stained by your Haematology departm
37. ulphatase 13 Iduronidase 13 Infantile Battens 13 Inhibin B itn erre eret RS 20 Insulin 45e eH REI EOM RA 20 K Krabbe ea iion CRINE EE Ee e d REUS 13 L Tet Ate costes eras tear rvrd Tet E i OE EHE 2 12 24 M MUERE I nn 20 Mannosidase P 13 Maninosidosis eese perse serena te Reden ae 13 Maroteaux Lamy 13 Metachromatic sse 4 13 Methionine loading test a9 Methylimalonate teh dps 19 MOV QUIO EI 13 Mucolipidosis 213 Mucopolysaccharidoses 13 Multiple hydrolases 13 Multiple sulphit ses 5 tr ER teer i inan 13 Mult ple sulphatidosis uie ehe 13 External Handbook This is a copy unless printed on controlled yellow paper Doc number CCL 003 Page 25 of 26 No unauthorized amendments or photocopies to be made Version number 1 5 Department of Chemical Pathology Great Ormond Street Hospital for Children N N Acetylaspartate eeeeeseeeeeeeeeeee nennen 19 N acetylglucosaminidase we 2l Neuraminidase 13 Neurodegenerative screen eese eese 14 Neuronal ceroid lipofuscinoses eee 13 Niemann Pick A amp B Non esterified fatty acids sse 12 O Organic acids eiecit ied tende pecie 12 19 Ornithine carbamoyl Transferas
38. umber type date and time of collection include medication diet fasting or fed sample name of sender address for report and invoice urgent contact name phone number if different from sender Labeling of Specimens Specimens should be legibly labeled with a minimum of three patient identifiers see above along with the date and time of collection type of specimen and specimen reference To avoid results being wrongly attributed to patients unlabelled samples or samples that do not match the name on the request form cannot be processed by the laboratory External Handbook This is a copy unless printed on controlled yellow paper Doc number CCL 003 Page 4 of 26 No unauthorized amendments or photocopies to be made Version number 1 5 Department of Chemical Pathology Great Ormond Street Hospital for Children Requirements for sample collection and processing are listed under individual analyses further on in this booklet Abbreviations used Li hep Lithium heparin L Liver Plain Plain container M Muscle RBC Erythrocytes F Fibroblasts WBC Leucocytes FB Fetal blood S Serum VL Vacuolated lymphocytes P Li hep plasma CV Chorionic villus B Whole blood CCV Cultured chorionic villus BS Blood spot AF Amniotic fluid Samples should be sent to us as soon as possible after collection However if storage is unavoidable guidance for sample storage is given under individual test Requestin
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