MTAP CLINICAL CHEMISTRY Types according to drainage characteristics:

 Blowout
CENTRIFUGE o Last drop of the liquid should be expelled into the
 Used to separate substances of different mass od density receiving vessel
 RPM – revolution per minute  Self-draining
 RCF – relative centrifugal force o Allow the contents of the pipette to drain by gravity
 RCF = RPM^2 x r x 1.12 x 10^-5
 RPM of the centrifuge is calibrated using tachometer Types according to purpose – measuring or graduating pipets
 Mohr pipette
Types of Centrifuge o Does not have graduations to the tip. It is a self-
 Horizontal head centrifuge draining pipette but the tip should not be allowed
o Swinging bucket type, the centrifuge tubes are held to touch the vessel while the pipette is draining
in a vertical position when not moving but are  Serological pipette
horizontal when the centrifuge is fully in motion o Has a graduation mark to the tip and is generally a
 Angle head centrifuge blowout pipette
o Has a fixed 25 – 52 degree at which the tubes are  Micropipette
held during centrifugation o Is a pipette with a total holding volume of less than
 Ultracentrifuge 1 ml
o Generates the highest speed; centrifuge head is
held at a fixed angle but generate tight sediment Types according to purpose – transfer pipettes
buttons due to the high speed generates  Ostwald Folin Pipette
o Have a bulb-like enlargement of the pipette stem
PIPETTES  Volumetric pipette
o Is designed to dispense or transfer aqueous solution
Types according to design: and is always self-draining
 To contain
o Holds a particular volume but does not dispense the Type or name function Drainage
exact volume Push button Deliver a Blow out
 To deliver or variable or last drop
o Will dispense the exact volume indicated micropipettes fixed volume
 Serological, Deliver a Blow out Principle:
standard type variable last drop  Light
volume o A form of electromagnetic energy that travels in
Kolmer Deliver a Drain to waves
serological variable baseline  Wavelength
volume
o Refers to distance between the peaks of a light
Mohr Deliver a Drain to
wave
variable baseline
o Is inversely proportional to amount of energy
volume
Capillary Contain a Wash out
fixed volume Beer’s Law
Lambda (two Contain a Wash out  Since path length (b) and absorptivity coefficient (a) are
types) fixed volume Blow out constants, we say that absorbance (A) is directly
Deliver a fixed proportional to the concentration (c)
volume
Ostwald-Folin Deliver a fixed Blow out
volume last drop
Volumetric, Deliver a fixed Drain by
standard type volume gravity
leaving last Components of spectrophotometer
drop in
pipette
 Light source
o For visible – infrared range: tungsten halogen
Cleaning of glassware
(iodide) lamp
 Presoaking glassware in soapy water is recommended
o For ultraviolet range: mercury arc lamp, xenon
 Cleaning solutions: potassium dichromate in H2SO4 or
lamp, deuterium discharge lamp
HNO3
 Monochromator
 Final rinses: type I or II water
o Examples of Monochromator: colored glass filters,
 Glassware are sterilized using dry oven (160-180C for 1 ½
prisms, interference filters, diffraction gratings
hour)
 Sample cuvette
o Holds the sample solution
PHOTOMETRY
o Can be plastic of glass
 Detector
 o Convert the transmitted radiant energy into an
equivalent amount of electrical energy
o Examples: photo cell, photo multiplier tube, photo
diode, barrier layer cell
 Readout system
o Measures the magnitude of the current generated
by the detector
o Galvanometer, ammeter
Variations of photometry
Fluorometry
Things to remember about Fluorometry:
 2 monochromators (dichroic beam splitter 1 and 2)
 Right angle
 UV light
 More sensitive/specific
QUALITY ASSURANCE
 Quality assurance / quality assessment
o Is a complete system of creating and following
procedures and policies to aim for providing the
most reliable patient laboratory results and to
minimize errors in the pre-analytical, analytical, and
post-analytical phases.
 Quality control
o Is an aspect of quality assessment that is used to
assess the analytical phase of patient testing
 Accreditation
o Is the process by which an agency or an
organization evaluates and recognizes a program of
study or an institution as meeting certain
predetermined qualifications or standards; applies
only to institutions and programs
Phases of analysis
 Pre-analytical phase
o Patient preparation
o Time of collection
o Specimen collection order
o Quality of specimen collected
o Specimen processing, storage and preservation
 Analytical phase
o Maintenance for equipment and instruments
o Calibration of equipment, verification of instrument
linearity
o Precision, accuracy and overall reliability check
through the use of standard materials, quality
control samples, procedures, and QC rules
o Accuracy – the nearness of closeness of the assayed
value to the true or target value
o Precision – the nearness of closeness of the assayed
value to a repeated value
o Repeatability – closeness of agreement between
results of successive instruments carried out under
the same conditions
o Reproducibility – closeness of agreement between
results of measurements performed under changed
conditions of measurements
o Practicability – the degree to which a method is
easily repeated
o Analytical errors
  Systematic errors (inaccuracy) – error that >500 mg/dl
occurs predictably once a pattern of Potassium <2.5 mEq/L
recognition is established; predictable >6.5 mEq/L
errors of the same sign and magnitude Bicarbonate <10 mEq/L
 Random errors (imprecision) – error that >40 mEq/L
occurs unpredictably; affects precision and Arterial of capillary <7.2
is the basis for varying differences between pH >7.6
repeated measurements Phosphate <1 mg/dl
Calcium <6 mg/dl
 Post-analytical phase
>13 mg/dl
o Delta check – checking the current results of a
Sodium <120 mEq/L
patient with his or her previous results
Arterial or capillary <40 mmHg
o Alarms and flags pO2
o Recording and reporting of results Arterial or capillary <20 mmHg
pCO2 >70 mmHg
Westgard QC rules
 1 2s – one control observation exceeding the mean +-2s. a Clinical chemistry conversion factors
warning rule that initiates testing of control data Analyte Conventional SI unit Conversion
 1 3s – one control observation exceeding the mean +-3s. unit factor
allows high sensitivity to random error Albumin g/dL g/L 10
 2 2s – two control observations consecutively exceeding the AST U/L (mU/mL) ukat/L 0.0167
same +2s or =2s. allows high sensitivity to systemic error Ammonia ug/dL umol/L 0.587
 R 4s – one control exceeding the +2s and another exceeding (NH3)
the -2s. allows detection of random error Bicarbonate mEq/L mmol/L 1.0
 4 1s – four consecutive control observations exceeding +1s (HCO3)
or -1s. this allows the detection of systemic error Bilirubin mg/dL umol/L 17.1
BUN BUN mg/dL 0.357
 10x – ten consecutive control observations falling on one
Calcium mg/dL mmol/L 0.25
side or the other of the mean (no requirement for SD size).
Chloride mEq/L mmol/L 1.0
This allows the detection of systemic error
Cholesterol mg/dL mmol/L 0.026
Cortisol ug/dL umol/L 0.0276
Clinical chemistry critical values Creatinine mg/dL umol/L 88.4
Bilirubin >18 mg/dl (newborn)
Glucose <40 mg/dl
 Creatinine mL/min mL/s 0.0167 o Cellulose
clearance o Chitin
Folic acid ng/mL Nmol/L 2.27
Glucose mg/dL mg/dL 0.0555 Must know
Hemoglobin g/dL g/L 10  Glycol aldehyde
Iron mg/dL umol/L 0.179 o Simplest carbohydrate (CHO)
Lithium mEq/L umol/L 1.0  Glucose, maltose, fructose, lactose, and galactose –
Magnesium mEq/L mmol/L 0.5 reducing substances/sugars
Osmolality mOsm/kg mmol/kg 1.0
 Sucrose – most common non-reducing sugar. Non-reducing
Phosphorus mg/dL mmol/L 0.323
sugar do not contain an active ketone or aldehyde group
Potassium mEq/L mmol/L 1.0
 The brain is completely dependent on blood glucose for
Sodium mEq/L mmol/L 1.0
Thyroxine ug/dL nmol/L 12.9 energy production – 2/3 of glucose utilization in resting
(T4) adults occurs in the CNS
Total g/dL g/L 10
protein Glycolysis Metabolism of
Triglycerides mg/dL mmol/L 0.0113 glucose to lactate or
Uric acid mg/dL mmol/L 0.0595 pyruvate
Vitamin B12 ng/mL pmol/L 0.0738 For production of
PCO2 / PO2 mmhg kPa 0.133 energy
Gluconeogenesis Formation of
Type of carbohydrates glucose-6-phosphate
from non-
 Monosaccharides
carbohydrate source
o Glucose
Glycogenolysis Breakdown of
o Fructose glycogen to glucose
o Galactose for use as energy
 Disaccharide Glycogenesis Conversion of
o Sucrose glucose to glycogen
o Lactose for storage
o Maltose Lipogenesis Conversion of
 Polysaccharide carbohydrates to
o Glycogen fatty acids
o Starch Lipolysis Decomposition of fat
Regulation of glucose metabolism METHODOLOGIES FOR GLUCOSE ASSAY
 Insulin
 Glucagon Chemical methods
 Epinephrine  Copper reduction
 Cortisol o Folin Wu method
 Growth hormone glucose
𝐶𝑢2+ → 𝐶𝑢1+
 Thyroxine reducing sugars
 Somatostatin 𝐶𝑢1+ + 𝑝ℎ𝑜𝑠𝑝ℎ𝑜𝑚𝑜𝑙𝑦𝑏𝑑𝑎𝑡𝑒 → 𝑝ℎ𝑜𝑠𝑝ℎ𝑜𝑚𝑜𝑙𝑦𝑏𝑑𝑒𝑚𝑢𝑛 𝑏𝑙𝑢𝑒

Specimen considerations and preparations o Nelson Somoygi

1+
 Whole blood, plasma, serum, CSF, urine, serous, synovial 𝐶𝑢 + 𝑎𝑟𝑠𝑒𝑛𝑜𝑚𝑜𝑙𝑦𝑏𝑑𝑎𝑡𝑒 → 𝑎𝑟𝑛𝑠𝑒𝑛𝑜𝑚𝑜𝑙𝑦𝑏𝑑𝑒𝑛𝑢𝑚 𝑏𝑙𝑢𝑒
fluid can be used as sample
o Neocupreine method
 Standard clinical specimen – fasting venous plasma 1+
𝐶𝑢 + 𝑛𝑒𝑜𝑐𝑢𝑟𝑝𝑟𝑒𝑖𝑛 → 𝑦𝑒𝑙𝑙𝑜𝑤 − 𝑦𝑒𝑙𝑙𝑜𝑤 𝑜𝑟𝑎𝑛𝑔𝑒 𝑐𝑜𝑙𝑜𝑟
 Fasting blood sugar should be obtained after 8-10 hours of
fasting
o Benedict’s method
 Whole blood gives 10-15% glucose levels than serum of
glucose
plasma 𝐶𝑢𝑆𝑂4
reducing sugars
 Serum is appropriate for glucose analysis if it is separated
→ 𝑔𝑟𝑒𝑒𝑛, 𝑦𝑒𝑙𝑙𝑜𝑤, 𝑜𝑟𝑎𝑛𝑔𝑒, 𝑏𝑟𝑖𝑐𝑘 𝑟𝑒𝑑 𝑝𝑟𝑒𝑐𝑖𝑝𝑖𝑡𝑎𝑡𝑒
from the cells within 30-60 minutes
 Glucose is metabolized at room temperature at a rate of o Ferric reduction
7mg/dL per hour (gray top) o Condensation method
 At 4C, glucose decreases by approximately 2mg/dL per hour Δ100 C
 2 mg of sodium fluoride per mL of whole blood prevents 𝑔𝑙𝑢𝑐𝑜𝑠𝑒 + 𝑜𝑟𝑡ℎ𝑜𝑡𝑜𝑙𝑢𝑖𝑑𝑖𝑛𝑒 𝑚𝑒𝑡ℎ𝑜𝑑
HAC
glycolysis for up to 18 hours → 𝑏𝑙𝑢𝑖𝑠ℎ 𝑔𝑟𝑒𝑒𝑛 𝑐𝑜𝑙𝑜𝑟
 Fluoride binds magnesium which causes inhibition of the
enzyme enolase Enzymatic methods
 CSF glucose concentration is approximately 60-70% that of  glucose oxidase
plasma concentration 𝑔𝑙𝑢𝑐𝑜𝑠𝑒 + 𝑜𝑥𝑦𝑔𝑒𝑛 𝑔𝑙𝑢𝑐𝑜𝑠𝑒 𝑜𝑥𝑖𝑑𝑎𝑠𝑒 → hydrogen peroxide
 CSF glucose should be obtained 1-2 hours before the spinal
tap o polarographic
 o colorimetric o Serves as the main storage for of lipid in man, as
ℎ𝑦𝑑𝑟𝑜𝑔𝑒𝑛 𝑝𝑒𝑟𝑜𝑥𝑖𝑑𝑎𝑠𝑒 + 𝑐ℎ𝑟𝑜𝑚𝑜𝑔𝑒𝑛𝑝𝑒𝑟𝑜𝑥𝑖𝑑𝑎𝑠𝑒 insulator or shock absorber, and as an integral part
→ water + oxidize chromogen of the cell membrane
 Cholesterol
 hexokinase-G6PD (oldest reference method) o Serves as part of cell membrane, as parent chain for
cholesterol based hormones such as aldosterone,
SPECIEM CONSIDERATIONS AND PREPARATIONS cortisol, and sex hormones
 Glucose in the solution exists either as an alpha-glucose or o Exists in the body in two different forms:
beta glucose.  Cholesterol ester – accounts to
o Alpha glucose is approximately 35% of total glucose approximately 70% of total cholesterol of
o Beta glucose is 65% the body, composed of a cholesterol ring
 Glucose oxidase is specific to beta with fatty acid
 Alpha glucose is converted into beta glucose using the  Free cholesterol – accounts to
enzyme mutarofase approximately 30% of total cholesterol; also
 Hemolyzed sample and IDKLOL cause decreased glucose known as unesterified cholesterol
with the Hexokinase-G6PD method
 Presence of strong reducing and oxidizing method Forms of lipids
 Phospholipids
LIPIDS o Structurally similar to triglyceride, except that 2
fatty acids and a phosphate group is attached to the
Functions: glycerol backbone
 Storage o Constituent of the cell membrane, contains polar
 Cushion and non-polar end
 Integral part  Free fatty acid
 Hormonal synthesis o Building blocks of lipids, hydrocarbon chains with a
terminal COO- group
Forms of lipids
TRANSPORT PROTEIN FOR LIPOPROTEINS – THE LIPOPROTEINS
 Triglyceride
o Possess 3 molecules of fatty acids and a molecule of  Apolipoproteins
glycerol which serves as the backbone
Functions Major source
Apo A-I Major Liver and Apo C-III Inhibits Liver
structural intestine lipoprotein
protein in HDL lipase
Activates LCAT Inhibits
– ligand for receptor
HDL binding recognition of
Apo A-II Structural Liver Apo E
protein in HDL Apo E2, 3, 4 Binds to LDL- Liver
Activated LCAT receptor and
Enhances remnant-
hepatic receptor
triglyceride Apo (a) Structural Liver
lipase activity protein for Lp
Apo A-IV Component of Intestine (a)
intestinal May inhibit
lipoproteins plasminogen
Apo B-48 Primarily Intestine binding
structural
protein in CHEMICAL COMPOSITION OF DIFFERENT LIPOPROTEIN CLASSES
chylomicrons Protein Chol Chol Trigly Phosphol
Apo B100 Major Liver (%) (%) ester (%) (%)
structural (%)
protein in CM 1-2 1-3 2-4 80-95 3-6
VLDL and LDL VLDL 6-10 4-8 16-22 45-65 15-20
Ligand for the
LDL receptor IDL interm ediat e bet ween vldl & ldl
Apo C-I Activates liver LDL 18-22 6-8 45-50 4-8 28-24
lipoprotein HDL 45-55 3-5 15-20 2-7 26-32
lipase
Apo C-II Activates Liver Separating lipoproteins
lipoprotein  Electrophoresis
lipase  Ultracentrifugation
Activates LCAT
Abnormal lipoproteins
  Beta-VLDL  Reagent: alcoholic potassium hydroxide
 Lp(a)  Purpose: cholesterol esters
o Step 2
 Reagent: petroleum ether
METHODOLOGIES FOR TRIGLYCERIDES DETERMINATION  Purpose: separate cholesterol from protein
o Step 3
 Enzymatic method  Reagent: acetic anhydride
o Initial enzymatic reactions: uses lipase and  Purpose: color reaction
glycerokinase
glucose SPECIMEN CONSIDERATIONS AND PATIENT PREPARATION
𝑇𝐴𝐺 → 𝑔𝑙𝑦𝑐𝑒𝑟𝑜𝑙 + 𝑓𝑎𝑡𝑡𝑦 𝑎𝑐𝑖𝑑𝑠
 Fasting requirement of at least 12 (to 14) hours
 No to Hemolyzed sample
glycerokinase  Avoid water contamination
𝑔𝑙𝑦𝑐𝑒𝑟𝑜𝑙 + 𝐴𝑇𝑃 → 𝑔𝑙𝑦𝑐𝑒𝑟𝑜𝑝ℎ𝑜𝑠𝑝ℎ𝑎𝑡𝑒 + 𝐴𝐷𝑃
 Serum or EEDTA-plasma may be used
 No to icteric specimens
 Chemical method – Van Handel Zilversmith
o Step 1
ELECTROPHORESIS
 Reagent: chloroform
 Regions are stained using Ponceau S, Coomasie brilliant
 Purpose: separate TAG and protein carrier
blue, amido black
o Step 2
 Electrophoretic patters:
 Reagent: alcoholic potassium hydroxide
o Beta-gamma bridging – seen in patients with liver
 Purpose: divide TAG
cirrhosis
o Step 3
o Bence Jones protein – seen in cases of monoclonal
 Reagent: sodium periodate
gammopathy
 Converts glycerol to formaldehyde
o ^ a2 microglobulin, v albumin – seen in patients
o Step 4
with nephrotic syndrome
 Reagent: chromotropic acid dissolve in
o v a-antitrypsin – seen in patients with emphysema
H2SO4
 serum is preferred; 24-hour urine and serous fluids can also
 Purpose: color reaction
be used
 no to lipemia
 Chemical method – Abell-Kendall
o Step 1  no to hemolysis
 Kjedhal method
o Reference method; quantifies protein by its
nitrogen content
o Assumes average nitrogen content of 16%
 Biuret reaction
o Measures protein by peptide bond
 Dyes
o Bromcresol purple – most sensitive, specific, and
precise among the dye-binding assays
o Bromcresol green – most commonly used
o Other dyes used: methyl orange, HABA
 Salt precipitation (turbidity) test – used for urine and CSF
MTAP CLINICAL CHEMISTRY DAY 2 o ATPase-dependent ion pumps

WATER BALANCE AND ELECTROLYTES Water – Osmolality

 Concentration of solutes per kilogram of solvent
Water – Introduction (millimoles/kg)
 40%-70% of body weight o Regulated by:
 Functions:  Thirst sensation
o Transport nutrients to the cells  Arginine vasopressin hormone (AVP)
o Determines cell volume  RAAS
o Removes waste products  ANP and GFR
o Act as body’s coolant o Thirst sensation
 Location:  Response to consume more fluids
o Intracellular fluid (ICF) – 2/3  Prevents water deficit
o Extracellular fluid (ECF) – 1/3 o Arginine vasopressin hormone (AVP)
 Antidiuretic hormone (ADH)
Distribution of Body Water in Adult  Increase reabsorption of water in kidneys
Compartment Percent of Percent of  Suppressed in excess H2O load
body weight total body  Activated in water deficit
water o Renin-angiotensin-aldosterone system (RAAS)
Extracellular  Liver  angiotensinogen (renin) 
Plasma 5 8 angiotensin I (ACE)  angiotensin II  AT1
Interstitial 15 25 receptor  kidney (sodium retention) /
Intracellular 40 67 adrenal gland (aldosterone release) /
Total body 60 100 vessels (constriction)
water o ANP
 Increase Na+ excretion in the kidney
The concentration of ions is maintained by: o GFR
 Passive Transport  Increase with volume expansion and
o Passive movement of ions across a membrane decrease with volume depletion
 Active Transport
o Required energy to move ions across the cell Electrolytes
membrane  Ions capable of carrying an electric charge
  Two types of ions: The Electrolytes
o Anion
 Ions that carry (-) charge and move toward SODIUM
the anode SODIUM – DESCRIPTION
 E.g. Cl-, HCO3, PO4  The most abundant cation in the ECF
o Cations  Major extracellular cation
 Ions that carry (+) charge and move toward  Na+, K+ -ATPase ion pump moves 3 Na+ ions out of the cell
the cathode in exchange for 2 K+ ions
 E.g. Na+, K+, Mg2+, Ca2+
SODIUM – REGULATION
Functions of Electrolytes  Plasma concentration depends in renal regulation
 Volume and osmotic regulation (Na+, Cl-, K+) o Intake of water
 Myocardial rhythm and contractility (K+, Mg2+, Ca2+)  Thirst
 Neuromuscular excitability (K+, Mg2, Ca2+) o Excretion of water
 Cofactors in enzyme activation (Mg2+, Ca2+, Zn2+)  AVP (increase H2O reabsorption)
 Regulation of ATPase ion pumps (Mg2+) o The blood volume status
 Acid-base balance (HCO3-, K+, Cl-)  Angiotensin II (increase aldosterone)
 Production and use of ATP from glucose (Mg2+ PO4-)  Aldosterone (increase Na+ reabsorption in
the kidneys)
Concentration of Cations and Anions in Extracellular and  ANP (increase urinary Na+ excretion)
Intracellular Water
Cation Extra Intra Anion Extra Intra SODIUM – CLINICAL APPLICATION
(mmol/L) (mmol/L) (mmol/L) (mmol/L)
Causes of hyponatremia (decreased Na+)
Na+ 136-145 15 HCO3- 23-29 10
Increase Na+ loss Increase H2O
K+ 3.5-5.1 150 Cl- 98-107 1
retention
Ca2+ 2.15-2.5 1 HPO42- 0.78- 50
1.42
Hypoaldosteronism Renal failure
Mg2+ 0.63-1 13.5 SO42- 0.5 10 Potassium deficiency SIADH
Diuretic use Nephrotic syndrome
Salt-losing Chronic heart failure
nephropathy
Severe burns Hepatic cirrhosis
Causes of hypernatremia (increased Na+) o Decreased function, decreased cellular entry
Excess Decreased H2O Increased intake or o Increased function, increased cellular entry
H2O loss intake retention
o Increased with exercise, diabetes mellitus, and cell
Diabetes Old/infant/mentally Cushing syndrome
insipidus breakdown
Profuse hyperaldosteronism
sweating POTASSIUM – CLINICAL APPLICATION
Severe Hypertonic salt
burns solution
Causes of hypokalemia (decreased K+)
GI loss Cellular shift – Renal loss Decreased
increase K+ intake
SODIUM – DETERMINATION uptake
 Specimen Vomiting Alkalosis Diuretics
o Serum, plasma (heparin and oxalate) (plasma)
Diarrhea Insulin overdose Renal tubular acidosis
o False increase with marked hemolysis Gastric suction Cushing’s syndrome
 Methods Laxatives hyperaldosteronism
o FES
o AAS Causes of hyperkalemia (increased K+)
o ISE (glass ion-exchange membrane) Decreased renal Increased Cellular shift Artifactual
excretion intake
Renal failure Oral/IV – K+ Acidosis Hemolysis,
POTASSIUM replacemen thrombocytosi
POTASSIUM – DESCRIPTION t s
 Major intracellular cation hypoaldosteronis Muscle/cellular injury Prolonged
o Regulation of neuromuscular excitability, m tourniquet
Chemotherapy/leukemi
contraction of heart, ICF volume, H+ concentration a
 Increased K+, increase cell excitability
(muscle weakness) POTASSIUM – REGULATION
 Decreased K+, increased cell excitability o Specimen
(arrhythmia or paralysis)  Serum, plasma (heparin)
 False increase with hemolysis
POTASSIUM – REGULATION  24-hour urine
 Aldosterone o Methods
o Increased K+ excretion in urine  FES
 Na+, K+ -ATPase pump  AAS
  ISE (use valinomycin membrane) o Schales and schales
o Colorimetric

CHLORIDE
CHLORIDE – DESCRIPTION
 Major extracellular anion
o Involve in maintaining osmolality blood volume and
electric neutrality (chloride shift)
o Rate limiting component in Na+ reabsorption
BICARBONATE
BICARBONATE – DESCTRIPTION
 2nd most abundant anion in the ECF
 Accounts for more than 80% of total CO2 with HCO3-
 A major component of the buffering system of the blood

CHLORIDE – CLINICAL APPLICATION BICARBONATE – CLINICAL APPLICATION

Increased HCO3- Decreased HCO3-
Metabolic alkalosis Metabolic acidosis
Severe vomiting Hyperventilation
Hypoventilation
Excessive alkali intake

BICARBONATE – DETERMINATION
 Specimen
o Serum, plasma (heparin)
o False decreased if left uncapped (decreased
6mmol/L per hour)
 Methods
o Enzyme method
CHLORIDE – DETERMINATION
 Phosphoenolpyruvate + HCO3 – PEP
 Specimen
carboxylase  oxaloacetate + H2PO4-
o Serum, plasma (lithium heparin)
oxaloacetate + NADH + H+ – MDH  malate
o False decrease with marked hemolysis (dilution)
+ NAD+
o 24-hour urine
 Methods
o ISE (use ion exchange membrane)
o Amperometric-coulometric (Cotlove Chloridometer)
MAGNESIUM Hyperthyroidism Digitales and digoxin
MAGNESIUM – DESCRIPTION Hypercalcemia
Diabetic ketoacidosis
 2nd major intracellular cation
 Neuromuscular conduction, enzyme cofactor and ATPase Causes of hypermagnesemia (increased Mg2+)
ion pump Decreased excretion, renal failure
 53% (bone), 46% (muscle, soft tissues), <1% (blood) Hypoparathyroidism
 In serum: 33% (protein bound), 61% (ionized), 5% Hypoaldosteronism
(complexed with PO4- and citrate) Increased intake (medications and therapy)
Bone carcinoma and bone metastases

MAGNESIUM – REGULATION CALCIUM

 Parathyroid hormone (PTH) – increased Mg2+ CALCIUM – DESCRIPTION
o Increased renal reabsorption and intestinal  For muscle contraction
absorption  For blood coagulation
 Aldosterone and thyroxine – decreased Mg2+  99% in bone and teeth and 1% in blood and ECF
o Promotes Na + renal reabsorption
o Increased renal excretion of Mg2+ and K+ CALCIUM – DISTRIBUTION
 Calcium in the blood is distributed as:
MAGNESIUM – CLINICAL APPLICATION o Ionized Ca+2
Causes of hypomagnesemia (decreased Mg+)  Unbound or free, physiologically active
Reduced intake Decreased absorption Others  45% of total calcium
Poor diet / starvation Malabsorption Excess lactation o Protein bound Ca+2
syndrome
Prolonged Mg+ - Diarrhea pregnancy  Bound to protein (e.g. albumin)
deficient IV  40%
Vomiting o Complex Ca+2
Laxative
 Bound to anions (e.g. HCO3-, PO4-, and
lactate)
Causes of hypomagnesemia (decreased Mg) – increased excretion
 15%
Renal Endocrine Drug induced
Tubular disease, Hyperparathyroidism Furosemide, thiazide
pyelonephritis
Glomeruloneohritis Hyperaldosteronism Gentamicin,
cyclosporine
CALCIUM – REGULATION  Methods
Factors affecting Ca+2 level in blood o AAS
 Bone resorption o ISE
o Cause increased Ca+2 in blood o Ortho-cresolphthalein complexone (CPC)
o PTH mobilizes Ca+2 from the bone o Alizarin dye, Arsenzo III dye, Methyl phenol blue
 Bone deposition o Clark and Collip (redox titration method)
o Cause decreased Ca+2 in blood o Ferro and Ham (precipitation with chloranilic acid)
o Calcitonin inhibits PTH and vitamin D
 Intestinal absorption PHOSPHATE
o Vitamin D increased Ca+2 in the intestine PHOSPHATE – DESCRIPTION
 Major intracellular anion
CALCIUM – CLINICAL APPLICATION  Component of phospholipids, nucleic acids, creatinine
Causes of hypocalcemia (decreased Ca+) phosphate and ATP
Hypoparathyroidism  80% bone, 20% soft tissues, 1% serum/plasma
Hypo/hypermagnesemia  GH decreased renal excretion of phosphate
Hypoalbuminemia
Acute pancreatitis PHOSPHATE – CLINICAL APPLICATION
Vitamin D deficiency Causes of hypophosphatemia (decreased PO4-)
rhabdomyolysis Hyperparathyroidism
Vitamin D deficiency
Causes of hypercalcemia (increased Ca+)
Hyperparathyroidism Causes of hyperphosphatemia (increased PO4-)
Malignancy (increased PTHrP) Increased intake
Increased vitamin D Increased released of cellular phosphate
Thiazide diuretics Increased breakdown of cells
Prolonged immobilization
PHOSPHATE – DETERMINATION
CALCIUM – DETERMINATION  Specimen
 Specimen o Serum, plasma (lithium heparin)
o Serum, plasma (dry lithium heparin) o 24-hour urine
o 24-hour urine o Hemolysis causes false increase
o Hemolysis causes false increase  Methods
 o Ammonium phosphomolybdate complex (340nm) o Increased in unmeasured cations
o Fiske-Subbarow method  Hypermagnesemia
 Hypercalcemia
LACTATE
LACTATE – DETERMINATION NON-PROTEIN NITROGEN
 Indicator of severity of oxygen deprivation (hypoxia)
 Liver converts lactate back to glucose “gluconeogenesis”

Lactate acidosis
Hypoxic conditions (type a) Metabolic origin (type b)
Lactate acidosis Diabetes mellitus, liver disease
Shock, MI, severe CHF Toxins (ethanol, methanol or
salicylate poisoning)
Pulmonary edema, severe
blood loss

ANION GAP
 Mathematical approximation of difference between the
concentration of unmeasured cations and unmeasured Compound Plasma conc. Urine conc.
anions (% of total NPN) (% of excreted N)
 (Na+) – (Cl- + HCO3-) Urea 45-50 86.0
 7-17 mEq/L Amino acids 25 ---
 Increased anion gap Uric acid 10 1.7
o Increased unmeasured anions Creatinine 5 4.5
 Uremia Creatine 1-2 ---
 Ketoacidosis Ammonia 0.2 2.8
 Lactic acidosis
o Increased in measured cations UREA
 Hypernatremia UREA – DESCRIPTION
 Decreased anion gap  The major waste product of the protein catabolism
o Decreased unmeasured anions  Excreted by the kidneys
 Hypoalbuminemia
Reference intervals UREA – PATHOPHYSIOLOGY
Adult Plasma/serum 6-20 mg/dL 2.1-7.1 Azotemia (increased urea in the blood)
mmol/L Prerenal Reduce blood flow of increased
Urine, 24-hour 12-20 g/day 11.5-4-4 protein catabolism
mmol Renal (uremia) Damage of nephrons (renal
urea/day failure/disease)
Postrenal Urinary tract obstruction
UREA – CLINICAL APPLICATION (Calculi, tumors)
Clinical application Conversion Decreased concentration
Renal function Urea N (mg/dL)  urea Low protein intake
(mg/dL) Severe vomiting and diarrhea
Hydration status Liver disease
Nitrogen balance 1 urea N  2.14 urea Pregnancy
Renal disease 0.467 urea  1 urea N
Adequacy of dialysis 0.357 /dL  mmol/L URIC ACID
URCI ACID – DESCRIPTION
UREA – DETERMINATION  Major end-product of purine catabolism
 Enzymatic method  Present as monosodium urates in plasma (98-100%
o GLDH coupled enzyme reabsorbed)
o Indicator dye  May precipitate in tissues (blood pH = 7; BUA > 6.8 mg/dL)
o Conductimetric (ISE) Reference intervals
 Chemical method mg/dL mmol/L
o Fearon’s reaction Adult male Plasma or serum 3.5 – 7.2 0.21 – 0.43
Adult female Plasma or serum 2.6 – 6.0 0.16 – 0.36
UREA – SPECIMEN CONSIDERATION Child Plasma or serum 2.0 – 5.5 0.12 – 0.33
 Use fasting blood Adult Urine / 24-hour 250-750 1.5 – 4.4
 Avoid fluoride or citrate anticoagulants mg/day mmol/day
 Refrigerate samples
URIC ACID – CLINICAL APPLICAITION  Increased triglycerides, decreased renal
Inherited disorders or purine metabolism urate excretion
Gout o Fructose intolerance
Renal calculi  Decreased F1PA
Uric acid nephropathy (chemotherapy)  Increased lactate, decreased renal urate
Kidney dysfunction excretion
o Hemolytic and proliferative process
URIC ACID – DETERMINATION  Increased metabolism of cell nuclei
 Enzymatic methods o Treatment of myeloproliferative disease with
o Spectrophotometric cytotoxic drugs
o Blauch and Koch  Increased destruction of cell
o Coupled enzyme (I) o Chronic renal disease
 Catalase  Decreased uric acid filtration and secretion
o Coupled enzyme (II) o Toxemia of pregnancy and lactic acidosis
 Peroxidase  Increased binding to renal tubules
 Chemical method o Purine-rich diet
o Phosphotungstic acid  Hypourecemia
o Caraway method o Decreased PRPP synthetase
 Decreased de novo purine synthesis
URIC ACID – SPECIAL CONSIDERATIONS o Liver disease
 Heparinized plasma, serum or urine  Decreased uric acid synthesis
 Lipemia, bilirubin, hemolysis (decrease UA) o Fanconi syndrome
 Salicylates and thiazides (increased UA)  Defective tubular reabsorption
o Chemotherapy with azathioprine or 6-
URIC ACID – PATHOPHYSIOLOGY mercaptopurine
 Hyperuricemia (enzyme deficiencies)  Decreased do novo purine synthesis
o Lesch-Nyhan syndrome o Overtreatment with allopurinol
 Decreased HGPRT  Decreased XO, decreased de novo purine
 Decreased reutilization of purine, increase synthesis
synthesis of purine
o Glycogen storage disease type 1
 Decreased G-6-P
CREATININE o Jaffe with absorbent
CREATININE – DESCRIPTION
 Chief product of muscle metabolism CREATININE – SPECIAL CONSIDERATIONS
 Not affected by protein diet  Falsely increase
o Glucose
Specimen Jaffe method o a-ketoacids
Unit mg/dL | umol/L o ascorbate
Adult male Plasma or serum 0.9 – 1.3 | 80 – 115 o uric acid
Adult female Plasma or serum 0.6 – 1.1 | 53 – 97 o cephalosporins
Child Plasma or serum 0.3 – 0.7 | 27 – 62 o dopamine
Adult male Urine 24-hour 800 – 2,000 mg/day  falsely decrease
Adult female Urine 24-hour 600 – 1,800 mg/day o bilirubin
BUN / Creatinine ratio: 10:1 to 20:1 o hemoglobin
o lipemic specimens
CREATININE – CLINICAL APPLICATION
 Sufficiency of kidney function CREATININE – PATHOPHYSIOLOGY
 Severity of kidney damage  increased concentration
 Progression of kidney disease o renal failure (glomerular function)
 Completeness of 24-hour urine o increased plasma concentration – decreased GFR

renal clearance and glomerular filtration rate
Glomerular filtration rate Volume of plasma filtered (V) AMMONIA
(creatinine clearance) by the glomeruli per unit of AMMONIA – DESCRIPTION
time (mL/minute)  By product of amino acid deamination
 Converted to urea in the liver
Specimen Reference value
ug/dL | umol/L
Adult Plasma 19 – 60 | 11 – 35
CREATININE – DETERMINATION
Child (10days-2yrs) Plasma 68 – 136 | 40 -
 Chemical method 80
o Jaffe reaction
o Jaffe kinetic

AMMONIA – CLINICAL APPLICATION
 Hepatic failure and hepatic coma
 Reye’s syndrome
 Inherited deficiencies of urea cycle
 Method
o Chemical method
 Ion-selective electrode
 Diffusion of H3 through selective
membrane into NH4Cl causing pH
change
 Spectrophotometric
 NH3 + bromphenol blue  blue dye
o Enzymatic method
 GLDH – decreased absorbance (340 nm)
 NH4+ + 2-oxyglutarate + NADPH +
H+ GLHD glutamate + NADO+
H2O
AMMONIA – SPECIAL CONSIDERATIONS
 Heparinized and EDTA tubes
 Samples are centrifuged at 0-4C within 20 minutes of
collection and the plasma/serum removed
 Avoid cigarette smoking for several hours
o Excretory (biliary) system
 Microscopic anatomy
o Lobules
Liver – Gross Anatomy
 Vascular system (two blood supplies)
o Hepatic artery (25%)
 Supplies oxygen rich blood
o Portal vein (75%)
 Supplies nutrient rich blood
 Excretory (biliary) system
o Excretory products of the cell drain to intrahepatic
ducts then to bile canaliculi
o Intrahepatic ducts  R + L hepatic ducts 
common hepatic duct + cystic duct  common bile
duct  duodenum
Liver – Microscopic Anatomy
 Lobules
o Functional units
o Six-sided with central vein and portal triads (hepatic
artery, portal vein, and a bile duct)
o Major cell types
 Hepatocytes and Kupffer Cells

LIVER FUNCTION TEST Liver – Biochemical Function

 Excretory and secretory
Liver o Eliminate heme waste products
 Largest internal organ  Hemoglobin is broken down:
 Gross anatomy  Globin, iron, heme
o Vascular system (two blood supplies) o Heme is converted to bilirubin and bound by
albumin (B1) then transported to the liver
  Synthesis Liver – Pathophysiology
o Synthesis and metabolism of carbohydrates Liver function alterations during disease
o Fat metabolisms  Jaundice (icterus)
o Plasma protein production o Yellow discoloration of skin, eyes, and
 Detoxification mucous membranes (>3 mg/dL)
o “first pass”  Pre-hepatic
o Foreign material (drugs and poisons) o Excessive destruction of
o Metabolic products (bilirubin and ammonia) RBC
o Acute and chronic
Liver – Determination hemolytic anemia
 Unconjugated bilirubin  Hemolytic disease
 Conjugated bilirubin of the newborn
 Delta bilirubin  Hemolytic
A. Measured using Diazotized Sulfanilic Acid transfusion reaction
 Direct Van den Bergh  Malaria – increased
o B2 + Diazo reagent  azobilirubin total bilirubin
B. Measures the Conjugated Bilirubin (increased B1,
 Indirect Van den Bergh normal B2)
o B1 + Diazo reagent  no reaction  Hepatic
o B1 + Accelerator + Diazo reagent  o Gilbert syndrome
Azobilirubin o Crigler-Najjar
Methods of Analysis o Dubin-Johnson
Malloy-Evelyn Jendrassik-Grof o Rotor syndrome
Accelerator 50% methanol Caffeine-Benzoate o Physiologic jaundice
Stopper None Ascorbic acid Total bilirubin B1 B2
pH Acidic (1.2) Basic (alkaline HDN, HTR Increased Increased Normal
tartrate) Gilberts syndrome Increased Increased Normal
End color Red-purple (560 Blue (600 nm) Crigler-Najjar syndrome Increased Increased decreased
nm)
B1 = total bilirubin – B2 Dubin Johnson Increased Normal Increased
Rotor Increased Normal Increased
Gall stones, tumors Increased Normal Increased
  y-globulins
 Post-hepatic  clotting factors
o Biliary obstructive disease
(cholestasis) Liver – Determination
o Increased total bilirubin  Kjeldahl
(normal B1, increase B2) o Nitrogen content
o Physical obstructions which  Refractometry
prevent flow of B2 into bile o refractive index
canal  Biuret
 Gall stones o Chelate (violet): Cu2+ and peptide bonds
 Tumors  Dye binding
 Cirrhosis o Spectral shift in the absorbance maximum
o Scar tissue replaces liver tissue resulting in  Electrophoresis
blockage of blood flow o Proteins separated based on electric charge
o Chronic alcoholism and chronic hepatitis C densities
 Tumors
o Primary or metastatic Liver – pathophysiology
o Benign or malignant  Albumin – decreased in chronic liver disease
o Reye’s syndrome  a1-globulins – decreased in a1-antitrypsin deficiency
o Disorders preceded by infectious (viral) or  y-globulin
drug (aspirin) related disease in children o increased in acute and chronic liver disease
 Drug and alcohol-related disorders o IgG and IgM – chronic active hepatitis
o Alcoholic fatty liver o IgM – primary biliary cirrhosis
o Alcoholic hepatitis o IgA – alcoholic cirrhosis
o Alcoholic cirrhosis  Clotting factors
 Drug-Related Disorder o Prothrombin time (PT)
o Acetaminophen  Increased PT in liver disease
o Inadequate production of clotting factors
Liver – Serum Protein  I, II V, IX, and X
 Albumin o Inadequate absorption of Vitamin K in the
 a1-globulins intestine
  Allosteric site
Liver – Enzymes o A cavity other than the active site that binds
1. Aspartate Aminotransferase (AST) regulatory (effector) molecules
2. Alanine Aminotransferase (ALT)
3. Alkaline Phosphatase (ALP) Enzymes – terms
4. Gamma Glutamyl Transferase (GGT)  Substrates
5. 5’-Nucleotidase (5NT)  Cofactors
6. Lactate Dehydrogenase  Isoenzymes
 Apoenzymes
 Holoenzymes
ENZYMES  Proenzyme or zymogens
 Allosteric enzymes
Enzymes – Description  Inhibitors
 Biologic proteins that catalyze biochemical reactions
 Not consumed or changed in composition Enzymes – Classification
 Found in all body tissue (intracellular) and is  Oxidoreductases
increased in serum after cell injury  Transferase
 Hydrolases
Enzymes – Function
 Lyases
 Hydration of carbon dioxide (respiration)
 Isomerases
 Nerve induction
 Ligases
 Muscle contraction
 Nutrient degradation (digestion) Enzymes – Mechanism
 Growth and reproduction  Lowering the activation energy
 Energy storage and use o Create an environment in which the
transition state is stabilized
Enzymes – Properties  Providing an alternative pathway
components of an Enzyme o Temporarily reacting with the substrate to
 Active site form an intermediate ES complex
o A cavity of an enzyme where substrates  Reducing the reaction entropy change
bind and undergo a chemical reaction
Enzyme – Mechanism  Liver enzymes
 Absolute specificity o ALT
 Group specificity o ALP
 Bond specificity o GGT
 Sterioisometric specificity  Pancreatic enzymes
o AMS
Factors that Affect Enzymatic Reaction o LPS
 Substrate concentration  Other enzymes
 Enzyme concentration o ACP
 Substrate and enzyme concentration o G-6-PDH
 pH
 temperature MI Profile Enzyme
 cofactors
 inhibitors CREATININE KINASE
MI – CK – DESCRIPTION
Enzymes – Determination  Storage of high-energy creatine phosphate in
 fixed time (two point) assay muscle cells
o reagents are combined and the amount of  Highest activities in skeletal muscle, heart (AMI),
reaction is measured and brain tissue
 continuous-monitoring or kinetic assays
o measurements at specific time intervals
o rate of change in substrate, cofactor,
product
o immunoassays MI – CK - DETERMINATION
o electrophoresis  Forward reaction (Tanzer-Givarg)
o Measure decreased in absorbance at 340
Enzymes – Clinical Significance nm
 MI profile o Optimum pH is 9.0
o CK  Reverse reaction (Oliver-Rosalki)
o AST o Increased in absorbance at 340 nm
o LDH o 6x faster than forward reaction
o Optimum pH: 6.8
MI – CK – SPECIAL CONSIDERATION MI – AST – DETERMINATION
 Source of error  Karmen method
o Hemolysis causes false increase due to AK o Uses malate dehydrogenase and monitors
activity decrease in absorbance at 340 nm
o CK is inactivated by light o Falsely increase in hemolyzed sample
o Physical activity and IM injections causes o Reference range: 5-30 U/L
increase CK
 Reference range LACTATE DEHYDROGENASE
o Male: 15-160 U/L MI – LDH – DESCRIPTION
o Female: 15-130 U/L  Interconversion of lactate and pyruvate
o CK-MB: <6% of total CK  Widely distributed highest activities in heart,
hepatic, skeletal muscle, and RBC
MI – CK – ISOENZYMES
Clinical significance
CK-3 / CK-MM / CK-2 / CK-MB / CK-1 / CK-BB / brain
muscle type hybrid type type
Slowest mobility 2nd fastest Migrate fastest
Major isoenzyme in Significant Highest
striated muscle and quantities in heart concentration in
normal serum tissues CNS, GI tract, and
MI – LDH – DETERMINATION
uterus (pregnancy)
 Wacker method
ASPARTATE AMINOTRANSFERACE o Forward reaction (lactate  pyruvate)
MI – AST – DESCRPTION o Increase in absorbance is monitored at 340
nm
 Serum glutamic-oxaloacetic transaminase (SGOT)
o Optimal pH is 8.3-8.9
 Transfer of amino group in aspartate and a-keto
 Wrobleuski La Due
acids
o Reverse reaction (pyruvate  lactate)
 Involved in the synthesis and degradation of AA
o Decrease in absorbance is monitored at 340
 Highest activities in cardia, liver, and skeletal muscle
nm
o Optimal pH is 7.1-7.4
MI – LDH – ISOENZYMES MI – MUST KNOW
Clinical significance CK-MB AST LDH
Isoenzyme Tissue Disorder (increase) Appearance 4-8 hours 6-8 hours 10-24 hours
LDH-1 (HHHH) Heart, RBC MI, hemolytic Peak 12-24 hours 24 hours 48-72 hours
LDH-2 (HHHM) anemia, RI, Stay elevated 3 days 5 days 10 days
megaloblastic
anemia LIVER ENZYME
LDH-3 (HHMM) Lung, spleen, Pulmonary  Alanine Aminotransferase (ALT)
pancreas embolism  Alkaline phosphatase (ALP)
LDH-4 (HMMM) Liver, skeletal Hepatic injury,  Lactate Dehydrogenase (LDH)
LDH-5 muscle skeletal muscle
 Gamma glutamyl transferase (GGT)
injury

ALANINE AMINOTRANSFERASE
MI – LDH – CLINICAL SIGNIFICANCE
LIVER – ALT – DESCRIPTION
 Relative concentration in normal serum
 Serum glutamic-pyruvic transaminase (SGPT)
 LDH-2 > LDH-1 > LDH-3 > LDH-4 > LDH-5
 Transfer of an amino group between alanine and
 In AMI and intravascular hemolysis, LDH-1 and LDH-
ketoglutarate
2 demonstrate a flipped pattern (LDH-1 > LDH-2)
 Increased in hepatocellular disorders

LIVER – ALT – DETERMINATION

Assay for enzyme activity
 Uses lactate dehydrogenase and monitors decrease
in absorbance at 340 nm
 Reference range: 6-37 U/L

ALKALINE PHOSPHATASE
LIVER – ALP – DESCRIPTION
 Catalyze the hydrolysis of phosphomonoesters
  Requires Mg2+ activator King and Armstrong Phenyl Phenol
 Evaluation of hepatobiliary and bone phosphate
Huggins & Talalay Phenolphthalein Phenol
diphosphate
Moss a-napthol a-napthol
phosphate
LIVER – ALP – ISOENZYME
 Liver ALP GAMMA GLUTAMYL TRANSFERASE
 Bone ALP LIVER – GGT – DESCRIPTION
 Placental ALP  Catalyze the transfer of the y-glutamyl residue from
 Intestinal ALP y-glutamyl peptides to amino acids, H2O
 Diagnosis hepatobiliary disorders and
LIVER – ALP – DETERMINATION
Assay for enzyme activity
 Bowers and McComb
 Cased on molar absorptivity of p-Nitrophenol
 Absorbance is measured at 405 nm LIVER – GGT – DETERMINATION
 Reference range: Assay for enzyme activity
o Adult: 30-90 U/L  Szaz Assay
o 0-3 months: 70-220 U/L o Absorbance of p-Nitroaniline is measured
o 3-10 years: 50-260 U/L 405-420 nm
o 10-puberty: 60-295 U/L
PANCREATIC ENZYMES
LIVER – ALP – DETERMINATION  Amylase (AMS)
Methods Substrate End product  Lipase (LPS)
Brodansky b-glycero- Inorganic PO4 +
Shinowara phosphate glycerol PE – AMS – DESCRIPTION
Jones  Catalyzes the breakdown of starch and glycogen via
Reinhart a, 1-6 branching linkages
Bessy, Lowry, & Brock p-nitrophenyl p-nitrophenol  Increased in acute pancreatitis
Bowers & McComb phosphate (yellow)
PE – AMS – DETERMINATION
  Amyloclastic o Formaldehyde and Cupric ions
 Chromogenic  Inhibits rec cell ACP
 Saccharogenic OE – ACP – DETERMINATION
 Continuous monitoring

PE – LP – DESCRIPTION
 Hydrolyzes of fats to produce alcohols and fatty
acids Methods Substrate
Quantitative end point Thymolpthalein
 Earliest and specific marker for acute pancreatitis
monophosphate
 Larger molecule remains in circulation (7 days)
Continuous monitoring a-napthyl phosphate

PE – LP – DETERMINATION
Cherry Crandall Tietz
Substrate 50% olive oil (triolein) 50% olive oil (triolein)
Titrating agent 0.4N NaOH 0.4N NaOH
Indicator Phenolphthalein Thymolpthalein +
veronal
Endpoint Fatty acid (oleic acid) Fatty acid (oleic acid)
End color Pink Blue

OTHER ENZYMES
 Acid phosphatase
 G-6-PDH

OE – ACP – DESCRIPTION
 Phosphatase inhibitors
o L-tartrate ions
 Inhibits specific prostatic ACP
 Total ACP – ACP after inhibition =
prostatic ACP