Transcript Assessment of Iron, Porphyrins, and Others

MLAB 2401: Clinical Chemistry
Keri Brophy-Martinez
Assessment of Iron, Porphyrins and Others
Laboratory Evaluation of Iron
Metabolism
Hematocrit
 Hemoglobin
 Red cell count + indices
 Total iron
 TIBC
 Percent saturation
 Transferrin
 Ferritin

Specimen Requirements:
Iron Studies
Serum without anticoagulant
– Plasma with heparin
–
– check product insert
– Oxalate, citrate or EDTA binds Fe ions, so
they are unacceptable
Early morning sample preferred due to
diurnal variation
– No hemolysis
–
Iron Panels

Three components
◦ Total serum iron
 Reflects the amount of ferric iron bound to
transferrin
 Does not include free iron found in hemoglobin
◦ TIBC= Total Iron Binding Capacity
 Measures capacity to bind iron with transferrin
 Sum of the serum iron and the UIBC
◦ Percent Iron Saturation/Transferrin
Saturation
 Ratio of serum iron to TIBC
Assessing Iron Levels and Forms

Directly measured
◦ Iron
◦ Transferrin
 Useful to diagnose hypochromic anemias
 Measured by the amount of iron it can bind
◦ Ferritin
 Best diagnostic test for IDA
 Reflects iron stores
 Acute phase reactant
Assessing Iron Levels and Forms
•
Indirect measure
– TIBC (Total iron-binding capacity)
• Measures the total amount of iron that
apotransferrin can bind
• TIBC= serum iron + unsaturated binding capacity
• Can be expressed as a percentage=percent
saturation
• Ratio of serum iron to TIBC
% Iron Saturation= (Total iron/TIBC) X 100
Test Methodologies: Iron
•
Colorimetric Procedure
– Separate Fe from transferrin with a strong
acid
– Iron is reduced from ferric(Fe3+) to
ferrous(Fe2+) state
– Addition of a chromogen creates a colored
compound
– Measurement of colored product by
spectrophotometry
Iron Reference Ranges
–
–
Diurnal variation
Men
 65-175 µg/dL
–
Women
 50-170 µg/dL
– Decreased Levels
• Decreased intake
• Increased need
• Increased loss
– Increased Levels
•
•
•
•
•
Increased absorption
Lead poisoning
Pernicious anemia
Megaloblastic anemia
Hepatitis
Test Methodologies:
TIBC & UIBC
Pre-treatment and Colorimetric Method
1. Add Fe3+ to saturate binding sites on
transferrin
2. MgCO3 is added to remove unbound
Fe3+
3. Mixture is centrifuged and the
supernatant tested using the serum iron
methodology
Reference Ranges

Transferrin
200-360 mg/dL

Ferritin
Male: 20-250 ng/mL
Female: 10-120 ng/mL

TIBC
250-425 µg/dL

% saturation
15-50%
Test Methodology: Porphyrins

Screening tests
◦ Urinary porphobilinogen (PBG)
◦ Urinary aminolevulinic acid (ALA)
◦ Urinary porphyrins

Quantitative Assays
◦ URO
◦ PROTO
◦ COPRO

Serve to classify porphyrias
Specimen Requirements:
Porphyrias
Protect specimens from light
 Urine samples

◦ Fresh
◦ Random
◦ 24-hour sample with no preservative

Blood samples
◦ Whole blood
◦ Plasma
Testing:
Porphyrias
◦ Ehrlich Watson test –outdated test to differentiate
between urobilinogen and porphobilinogen.
 Urobilinogen is extractable into either chloroform or
butanol.
 Porphobilinogen is neither, and is left behind.
◦ Hoesch rapid screening test for porphobilinogen
◦ Fluorescent technique for other porphyrins.
◦ Mauzerall-Granick–more reliable screening test
Lab Methods: Myoglobin
Procedures incorporate the binding of
specific antibodies to myoglobin with a
resulting chemical or physical change that
can be measured and correlated to
myoglobin concentration
 Specimen requirements

◦ Usually plasma
 check product insert
Specimen Requirements: Lead

Whole blood
◦ Why? Circulating lead found in the RBC
 Royal blue top with EDTA anticoagulant
 EDTA
◦ Lead-free containers
◦ Venous sample preferred
 Capillary specimens can be used for screening

Urine
Lab Method: Lead

Test methodologies
◦ AAS
◦ Anodic stripping voltammetry

Reference Ranges in blood
Children: < 10 µg/dL
Adult :<25 µg/dL
References



Bishop, M., Fody, E., & Schoeff, l. (2010). Clinical Chemistry:
Techniques, principles, Correlations. Baltimore: Wolters
Kluwer Lippincott Williams & Wilkins.
http://www.clsi.org/source/orders/free/h17-a.pdf
Sunheimer, R., & Graves, L. (2010). Clinical Laboratory
Chemistry. Upper Saddle River: Pearson .