1. INTRODUCTION TO CHEMICAL PATHOLOGY
Prof. David Marais
6.33 Falmouth Building
UCT Health Science Faculty & NHLS
Anzio Rd, Observatory, 7925, South Africa
Ms Farhana Hassan - Telephone O21 4066192
UCT MBChB Semester 3 Language of Medicine
Wednesday 16th January 2013, 09:00-10:00
New Learning Centre
2. Aqenda
Chemical Pathology
Division of Chemical Pathology
Chemical Pathology Course
2 lnvestigations
Methods
lnterpretation
lmplications
3 Chemical Pathology and Medical Practice
Systems approach and problems
3. Definition of Chemical Patholoqv
SYNONYMS ClinicalBiochemistry,ClinicalChemistry
Pathologic Biochemistry
(Metabolic Medicine)
ETYMOLOGY Pathology: knowledge & study of disease
Chemistry: knowledge and study of composition and
properties and reactions of substances;
Biochemistry: chemistry pertaining to life
Anatomical Pathology: macro- & microscopic apearance
Medical Microbiology: disease from microorganisms
Forensic Pathology: causes of death, natural/criminal
4. FORMAL DEFINITION OF CHEMICAL PATHOLOGY
A discipline in the Health Sciences concerned with the description of the
(bio)chemical constitution of humans evidenced by samples during human
Health and Disease, in relation to environmental or genetic factors or
metabolic stress. The changes vary in scope and degree, signifying success
of homeostasis and prognosis. The findings are used in the management of
patients, in detecting unfavourable changes in health ahead of disease so as
to anticipate and avoid or ameliorate disease, and to gain insight into the
mechanisms of disease. Like other disciplines in healthcare the
commitment is to the patient care.
ln the modern era places more reliance on objective description of disease;
in 75% of medical diagnoses a laboratory test is required. This discipline
requires interaction with clinical practitioners (contributions to special clinics
are increasing), other disciplines in pathology and (bio)chemistry as well as
genetics. Like other disciplines in Healthcare, chemical pathology not only
provides service but has special requirements for its practice, researches the
pathophysiology of disease and diagnostic strategies whilst being aware of
financial resources. Subspecialties are emerging as technology and clinical
demands increase
5. ORGANISATION OF CHEM PATHOLOGY
CLINICAL LABORATORY SCIENCES
1. Anatornical Pathology
2. Chemical Pathology
3. Forensic Pathology
4. Haematology
5. Human Genetics
6. lmmunology
7. lvedical Biochemistry
8. lVicrobiology
CHEM ICAL PATHOLOGY
(UCT & National Health Laboratory Service)
Falmouth Building: Offices and Research Laboratories
Prof. Marais, Dr Blackhurst, Dr King, Ms Leisegang, Dr Mcca(hy, Mr Mohamed,
Ms Solomon,Ms Ratanjee, Mr Woolley. Students.
Groote Schuur Hospital C17: Clinical Service Laboratory
Dr Haarburger, Dr King, Dr Omar, Dr Vreede. Dr Fortgens. Registrars.
Red Cross Hospital ICH: Clinical Service Laboratory, Metabolic Disorders
Dr G vd Watt
6. CHEMICAL PATHOLOGY COURSE
. Lectures, tutorials, patientbased problems
a Reference books, in-house notes
. Principles & Concepts > facts
. Scientific vs Clinical Approach (assumptions, general experience)
. Emphasise Diagnosis & Disorders rather than Chemistry
. Read & Understand, distill to principles and vital facts
. Revise anatomy biochemistry, chemistry, physiology
7. STUDY MATERIAL
Marshall and Bangert. Clinical Chemistry,
6th ed. Published in 2008 by Elsevier -
CHEMICAL
ISBN'l 0:0-7234-3455-7 PATHOLOGY
I,TiCTI]RE NOTES B
J. Baynes & Marek Dominicza k
Medical
Biochemistry, Mosby 2nd Ed. 2005
Paula Yurkanis Bruice. Essential Organic
Chemistry, Pearson lnternational Edition or
2nd Edition, Pearson.
Silberberg, (McGraw Hill) Chemistry. The
Molecular Nature of Matter and Change 3rd,
4th or 5th edition, Mccraw Hill
Other Text Books. Journals
Especially reviews
lnternet search
Discussions
w
8. CHEMICAL PATHOLOGY SERVICE
IN DICATIONS
DIAGNOSIS: Differentialdiagnosis(chestpain)
SCREEN: Disorders without much clinical manifestation (thyroid)
MON ITOR: Response to treatment (hypokalaemia, diabetes)
PROGNOSIS: Susceptibility to disease or complications (cholesterol)
Key tests and Related tests in context
(heart failure electrolytes, renal function, thiamine deficiency)
SAM PLES
Blood (venous/arterial), serum or plasma. Phlebotomists.
U rine (dipstick or test tube analysis)
Stool (occult blood, fat malabsorption by stool collection)
CSF (glucose, protein)
Effusions (protein)
9. SAMPLE ANALYSIS
CONSENT Explanation. Verbal consent unless invasive.
PREPARATION Fasting,provocativetests.
TIMING Diurnal rhythms.
PRESERVATION Appropriateforanalysis
Blood: Heparin or EDTA anticoag, NaF stops glycolysis
Gases escape. Gln degradation to NH3
Urine: toluene or azide to prevent bacterial growth
acidify for Mg, Ca, alkalinise for urate
TECHNIQUE Venesection, etc. Stasis, haemolysis, bleed into...
LABELLING Medicolegal ! Leave sample visisble, tube managable
INFORMATION Report in relation to result, associated tests
RECEPTION On time for lab service !
COST Limit number. Preliminary test before additional.
PROBLEMS
Pre-pre-analytical Preparation,posture,technique,forms
Pre-Analytical Transport, separation, aliquots, 2nd container & store
Analytical Lab quality control.
PostAnalytical Report and lnterpretation. Delay to attention.
10. METHODS
Manual or Automated Analysis.
Analyte in matrix, interfering substances.
lnspection - turbidity, lipaemia, haemolysis, colour, odour,
COLORIMETR IC
Chemical reaction coupled to colour product, direcuindirect
Short times, inexpensive, automatable, not very sensitive (mmol/L to FimoUL)
U rea, creatinine, albumin
FLU OR IMETRIC
Molecule or product responds to Excitation l, Emission.l.
Not as quantitative but more sensitive (pmol/L to nmol/L).
Manual, seldom in routine labs.
ION SENSE ELECTRODES
Selective permeability to ion changes.
lnexpensive, rapid, automatable, measures unbound ions.
H. for pH, Na., Ca..
11. METHODS
ENZYMATIC ASSAYS
Enzyme specific for substrate yields product that can yield indicator or
cofactor can indicate reaction
Lactate dehydrogenase uses NAD and generates NADH (UV absorbance)
Lactate + NAD. +pyruvate+NADH +H.
Concentration: molar extinction coefficient, standards
Activity: moles/time (units) relate to volume; for enzyme description.
IMMU O-ASSAYS
N
Antibodies (Ab) are highly specific to epitopes on protein
Analyte = antigen (Ag). Can be radio labelled and added to reaction (Ag*)
Very sensitive, specific, some automation, costly to produce Ag and Ab
Competitive assay: add Ag*, mix with Ag
Add specific antibody (Ab) to bind Ag and/or Ag* and form complex
Analyse radio-activity in complex
No Ag means only Ag* binds and radioactivity is high
Large amount Ag successfully competes against Ag*, counts are low
12. METHODS
ELECTROPHORES IS
Proteins in buffer (sustains charge) will migrate (to anode).
Agarose migration more complex, acrylamide by size
Stain, patterns: monoclonal lg, lsoenzymes
Relatively inexpensive, sensitive
CH ROMATOG RAPH Y
Separation between mobile and stationary phases
Thin Layer Chromatography (TLC): simple, low cost, time, qualitative
lon Exchange Chromatography: amino acids
Gas Chromatography (volatile by derivative): organic acids, sterols, fatty acids
DNA
Polymerase chain reaction (PCR); amplifies DNA (blood, buccal)
Generally for specific mutations by several techniques
Biochemical phenotype similar from different genes, also polymorphisms
Founder effects more cost-effective
13. INTERPRETATION
IDENTITY Ensure correct sample, time
CONTEXT Symptoms,signs,treatment,differentialdiagnosis
Can assay assist in solving problem ?
Pitfalls in interpretation (T4 & TSH on treatment)
UNITS Reference range (age, gender, ...)
Report in Sl vs other units. (TC 5.18 mmo/L = 200m9/dL)
1 mole = atomic (molecular) mass in grammes. (Avogadro's Nr)
NaCl: 23 da + 35 da = 58 da. Thus 589 in 1L is 1 mol/L
Plasma concentration of Na. is -140 mmol/L
"Physiologic saline" is 0.99/dL or 155mmol/L (31omosm/L)
mmol/L Na., K., Ca.., M9.., urea, glucose, triglyceride, cholesterol
pmol/L creatinine, iron, albumin (-600, or 409/L)
nmol/L cortisol (-500), H. (40, or pH= 7.40),
pmol/LACTH (50)
15. STATISTICS
DESCRIPTIVE Mean (avg) of observations (>) = ( L / n) {NB: n-1 for sample)
Median (midway in observations)
Variance=r(r_obs),
Std deviation = ., Variance
Std error of mean = StdDev / (., n)
CV=StdDev/x
Reference lnterval (normal ranoe): aoe. oender. socio-economic. oenes
Generally >120 samples, well collected and analysed
Centiles along range, lowest end is 1 and highest 100
Precision = closeness of repeated observations
Accuracy = conformance with exact standard
Sensitivity (%) = proportion with disease who test +ve
Specificity (%) = proportion without disease who test -ve
CHAN GES Cvtotd of parameter relates to CV""""y & Cvbiol
TC: Cvbior 5% and
-Cv""""r
3%
CV,*",= [(5)'+(3)']= [34] =5.8
Thus on one repeat test require a change of > 11o/o
16. IMPLICATIONS OF INVESTIGATION
INSIGHT Need for test, implications for management.
Confidentiality.
lnformed consent for H lV disease.
Gate-keeping of requests.
U RGEN CY Relates to clinical decision making and cost
Turn-Around-Time in lab is geared to this.
Critical values are generally reported to Dr (N B contact!)
Emergency: treat hypoglycaemia regardless
DOCU MENT Sending and Receiving
Response to result
17. COMMON CLINICAL SETTINGS FOR INVESTIGATION
Acid/base disturbances GIT disturbances
acidosis/alkalosis diarrhoea
respi ratory/m etabolic malabsorption
Water and electrolyte disturbances pancreatitis
overhydration & dehydration malignancy
hypematraemia & hyponatraemia Endocrine disease
hyperkalaemia & hypokalaemia diabetes
Calcirm and phosphate disorders thyroid
bone disease parathyroid
endocrine disorders Metabolic disease
Carbolrydrate homeostasis u ric acid (gout)
diabetes and hypoglycaemia iron metabolism
Liver dysfunction dyslipidaemias
hepatitis-cirrhosis€ilu re inherited metabolic disease
metabolic disorders nutritional (vitamins)
jaundice (bili,transaminase,alk phos) Neoplastic disease
Kidney dysfunction tumor markers (PSA, CEA)
acute vs chronic failure protein (myeloma)
nephrotic syndrome Toxicology
metabolic disorders paracetamol
Heart disease aspirin,organophosphate, ...
acute myocardial infarction (CK) alcohol
Fe metabolism, thiamine, ...
18. CONCLUSIONS ABOUT CHEM PATHOLOGY
1. Vital for diagnosis & management of many disorders and thus has
serious responsibilities in healthcare
2. Appropriate investigation for patient requires good clinical assessment
and careful consideration of tests (likelihood, differential, cost)
3. Responsibility of requesting doctor to ensure results are applied
4. Must be understood and be related to anatomy, physiology, chemistry,
biochemistry, pathology, microbiology, pharmacology, ...and all else!
5. Application is not restricted to laboratories but open to all medical
practitioners as sideroom investigations (urine dipstick) and pointof-
care testing.
6. ls a fascinating discipline which will still improve our insights into disease,
its diagnosis and its treatment in future.
