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HBC/BC 306

Clinical Biochemistry

Basic biochemistry emphasizing human

metabolic pathways & their relationship

to health & disease.

Biochemical tests are used in the

diagnosis, management, treatment and

subsequent follow-up, of human disease

Type of Samples

Concentration of different analytes determined

in:

BloodSerum or plasma

Urine

Cerebrospinal fluid (CSF)

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Within-Individual Variation

1. Time of day- diurnal variation of:

Plasma iron

Adrenocorticotropic hormone (ACTH)

Cortisol

2. Diet

Plasma [triglyceride]

Response to glucose tolerance tests

Urinary calcium excretion

3. Muscular Exercise

Increase plasma creatine kinase activity

Increase blood [lactate]

Lower blood pyruvate

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4. Menstrual Cycle

Plasma [iron]

Plasma conc. of pituitary gonadotrophins

& ovarian steroids & their metabolites

Amts of hormones & their metabolites

excreted in urine

5. Drugs

Oestrogen-containing oral contracetives

affect plasma constituents

Between-Individual Variation

1. Age

Plasma [phosphate] & alkalinephosphatase activity

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Plasma & urinary conc. of gonadotrophins

& sex hormones

2. Sex

Plasma creatine, iron, urea conc.

Plasma & urinary conc. of sex hormones

3. Race

Plasma [cholesterol] & [protein]

Diet???

Reference Ranges

Set of results from a particular defined

population

No clear-cut distinction between normal &

abnormal conc. of any constituent

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To interpret results, know:

Reference range for healthy individuals

Expected values for patients with disease

Prevalence of disease in population

Sensitivity of test

Ability to show +ve results in patients with

particular disease (true +ve rate)

The higher the sensitivity, the greater the

detection rate, the lower the false ve rate

Specificity of test

% of ve results among people who do not

have the disease

The higher the specificity, the lower the

false +ve rate

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Use of Enzymes in Clinical Biochemistry

1. Indicators of various diseases

2. Analytical reagents

3. Therapeutic agents

Release of Enzymes from Cells

1. Necrosis or severe damage to cells

2. Increased rate of cell turnover

3. Increased conc. of enzymes within cells

4. Duct obstruction

Enzymes Commonly Used in Diagnosis

1. Lactate dehydrogenase (LD)

2. Creatine kinase (CK)

3. Transaminases

4. Alkaline phosphate (ALP)

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Serum Markers in the Diagnosis of

Tissue Damage

Myocardial Infarction

Myocardial infarct - lesion formed when

cells of the myocardium die due to severe

ischemia (territorial distribution of blood)

Common cause- Artery obstruction caused

by formation of thrombus (blood clot)

Anti-thrombolytic therapy- protects

myocardium from permanent damage by

restoring blood flow

Streptokinase

Recombinant tissue plasminogen

activator

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Measurement of circulatory proteins

(enzymes & non-enzyme proteins)

released from necrotic myocardial

tissue are useful in diagnosis

Rise rapidly to a peak between 18 &

36 hrs

Return to normal dependent on life

each protein in the plasma

1. Myoglobin

earliest marker

not cardiac specific

raised by any form muscle damage

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2. Total CK

3 principal CK isoenzymes

Dimers BB, MB, MM

Rel. early marker

Not cardiac specific

3. CKMB

Rel. early marker

Higher cardiac specificity over totalCK

4. Cardiac Troponin T

Rel. early marker

Cardiac specific

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But elevated in diseases of

regeneration of skeletal muscle &

chronic renal disease

5. Cardiac Troponin I

Rel. early marker

Highly cardiac specific

6. LDH Isoenzymes

Late marker

Not cardiac specific

LD1/LD2 determination increases

cardiac specificity

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Acute Pancreatitis

Obstruction of pancreatic duct that delivers

pancreatic juice to small intestine

Gall stones

Alcohol abuse

Inappropriate release of pancreatic enzymes

& their premature activation

Eg Trypsinogen is activated to trypsin

- converts many other enzymes to

their active forms

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Lab. Diagnosis involves measurement of

pancreatic digestive enzymes.

1. Serum Amylase

Elevated levels sensitive diagnostic

indicator Low specificity

Many non-pancreatic causes of

hyperamylasemia

2. Serum Lipase

Higher specificity than amylase

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Renal Function Tests

To identify renal dysfunction

To diagnose renal disease & monitor

progress

To monitor response to treatment

Functions of the Kidney

1. Production of urine

Elimination of metabolic end products,

ie urea & creatinine Elimination of foreign materials drugs

Control of volume & composition of

extracellular fluid

- water & electrolyte balance

- acid / base balance

2. Endocrine Functions

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- Vitamin D

- Erythropoietin

- Antidiuretic hormone

Biochemical Tests

- Urinalysis

- Measurement of glomerular filtration

rate

- Tubular function tests

Tests rarely establish the cause BUT help in

screening for damage & monitoring progression

of disease

Urinalysis

1.Valid sample = fresh sample

2.Appearance- Unusual colour due to blood or

infection?

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3.Specific gravity- sticks measure ionic

species only & NOT glucose

4.pH normally acidic except after a meal

5.Glucose increased blood glucose??

6.Proteinuria- detected using urine sticks

- Raised plasma low MW proteins, Bence

Jones, myoglobin?

- Glomerular leak?

- Decreased tubular reabsorption of

protein?

7.Urine sediments- microscopic examination

for cells, fat droplets?

Measurement of Glomerular Filtration Rate

(GFR)

- GFR is essential to renal function & is a

frequently performed test

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- Measurement is based on concept of

clearance

The determination of the volume of plasma

from which a substance is removed by

glomerular filtration during its passage

through the kidney

Clearance = (U x V) / P

Where U = urinary conc. of substance

V = rate of urine formation in ml/minP = plasma conc. of substance

Units = vol / unit time (ml/min)

Eg. Creatinine Clearance (ml/min)

=Urine[creatinine] x urine vol (ml) Plasma [creatinine] collection time (min)

If clearance = GFR then substance

- Freely filtered by glomerulus

- Glomerulus is sole route of excretion, ie

no tubular secretion or reabsorption- Easily measurable

- Has constant plasma conc.

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Tubular Function Tests

Specific renal tubule disorders may affect ability

to

Concentrate urine

Excrete appropriately acidic urine

Cause impaired reabsorption of phosphate,

amino acids, glucose

Acute Renal Failure (ARF)

Due to reduced glomerular filtration rate with

resultant retention of

Urea & creatinine

Na & water

Acid with metabolic acidosis

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Potassium with hyperkalaemia (potassium is

released from cells & acidosis promotes the

leakage)

ARF may manifest as pre-renal, renal or post-

renal

Chronic Renal Failure (CRF)

Progressive loss of nephrons resulting

permanently impaired renal function

Retention of urea, creatinine, toxins

Rate of urea excretion falls & cannot

balance rate of production

Plasma urea rises, urea conc. in filtrate of

functioning nephrons rises

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May cause osmotic diuresis in these

nephrons

Sodium & potassium excretion?

Liver Function Tests

Functions of the Liver

1. Carbohydrate metabolism

Glycogen synthesis, storage &

breakdown

Gluconeogenesis

2. Lipid metabolism

Synthesis of almost all lipoproteins,phospholipids, cholesterol

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3. Protein Synthesis

Many plasma proteins, most coagulation

factors are synthesized in hepatic cells

4. Storage functions

Vitamins A, D, B12 & iron

5. Excretion & Detoxification

Bile pigments & cholesterol are excreted

in bile

Many drugs are detoxicated by the liver

Ammonia derived from amino acid

metabolism is converted to urea

Steroid hormones are inactivated byconjugation with glucuronate & sulphate

in liver & excreted in urine

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6. Reticulo-endothelial function

Kupffer cells lining sinusoids of liver form

part of reticulo-endothelial system

Bilirubin Metabolism

Heme groups (from haemoglobin

myoglobin, cytochromes) are freed of iron

& converted to bilirubin

Bilirubin is a yellow pigment that is

strongly lipophilic & cytotoxic

Transported to liver, bound to serum

albumin

At the liver, bilirubin is coupled to

glucuronic acid & the conjugated bilirubin

is excreted into bile

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Jaundice

Clinically detectable increase in plasma

bilirubin levels

Due to predominant rise in unconjugated

bilirubin from excessive haemolysis or

decreased excretion

Biochemical Tests in Liver Disease

Hepatocellular Damage

Damage to liver cells, with or without

necrosis, causes acute release of intracellular

constituents into bloodstream

Detected by measuring plasma enzymes,

alanine & aspartate aminotransferases

(transaminases)

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Biliary Tract Involvement

Characterized by increased production &

raised plasma levels of hepatobiliary

enzymes, alkaline phosphatase (ALP)&

gamma- glutamyltransferase (GGT)

Obstruction of biliary tract with jaundice

indicates cholestasis

Impaired Hepatocellular Function

Affects synthesis of albumin, coagulation

factors & bilirubin metabolism

Hypoalbuminaemia & a prolonged

prothrombin time are detectable when

damage is extensive & prolonged

Deficiency of Vit K may also cause

prolonged prothrombin time

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Disordered Metabolism

Patients with severe liver disease may have:

Significant decreases in plasma [urea] due to

failure of liver to convert amino acids &

ammonia to urea

Hypoglycaemia due to impaired

gluconeogenesis or glycogen breakdown

Raised conc. of all plasma lipid fractions, if

cholestasis is present

Specific Liver Diseases

Acute Hepatitis

Caused by viruses or toxins

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Cell damage detectable by plasma enzyme

estimations

Cirrhosis

During active cellular destruction,

transaminase levels rise, sometimes with

jaundice

In alcoholic cirrhosis, GGT levels are

elevated