CLINICAL CHEMISTRY

BY: DON ALGHERNON T. CHENG Quality Control

Conversion factors  A system of ensuring accuracy and precision in the
laboratory by including quality control reagents in every
Albumin, Total Protein 10
series of measurements.
Bilirubin 17.1
Cholesterol 0.0264  A process of ensuring that analytical results are correct by
Creatinine 88.4 testing known samples (control solutions) that resemble
Thyroxine 12.9 patient samples.
Glucose 0.055  It involves the process of monitoring the characteristics of
Calcium 0.25 the analytical processes and detects analytical errors during
Magnesium 0.5 testing.
BUN 0.357
Uric Acid 0.0595
1. Sensitivity – Is the ability of an analytical method to measure
Creatinine clearance 0.0167
the smallest concentration of the analyte of interest.
Iron 0.179
2. Specificity – Is the ability ofan analytical method to measure
only the analyte of interest.
Seven Base SI units 3. Accuracy – The nearness or closeness of the assayed value to
the true or target value.
a. meter (m)= length
4. Precision or reproducibility – The ability of an analytical
b. kilogram (kg) = mass
method to give repeated results on the same sample that agree
c. second (s)= time
with one another.
d. mole (mol) = quantity of substance
5. Practicability – The degree by which a method is easily
e. ampere (A) = electric current
repeated.
f. Kelvin (K) = thermodynamic temperature
6. Reliability – The ability of an analytical method to maintain
g. candela (cd) = luminous intensity
accuracy and precision over extended period of time during
* Enzyme Unit =U/L or katal unit
which equipment, reagents and personnel may change.

Types of Error
1. Random Error
 Is present in all measurements; it is due to chance.

DON ALGHERNON T. CHENG, RMT 1

  It us due to instrument, operator and blood ratio
environmental conditions (variations in techniques) 7. improper mixing
Ex. Pipeting errors, mislabeling samples, of sample and
additives
temperature fluctuation, improper mixing of sample
8. incorrect
and reagent.
specimen
2. Systematic Error preservation
 Is an error that influences observations consistently 9.Incorrect used of
in one direction (constant difference) tubes from blood
Ex. Calibration problems, deterioration of reagents collection
and control materials, unstable and inadequate 10. Mishandled
specimen ( transport
reagents blanks, contaminated solutions, failing
and storage)
instrumentation and poorly written procedures. 11. Missed or
3. Clerical Error incorrectly
 The highest frequency of clerical errors occurs with interpreted
the use of handwritten labels and request forms. laboratory requests.

Statistics

1. Mean – a measure of central tendency

Pre- analytical Errors Analytical Errors Post analytical Errors
2. Standard Deviation (SD) – a measure of the dispersion of
1. Improper patient *Errors happened 1.Unavailable or
preparation due to improper delayed laboratory values from the mean.
2. Mislabeled machine calibration results. 3. Coefficient of Variation (CV) – a percentile expression of
specimen or wrong processes 2. Incomplete the mean.
3. Incorrect order of laboratory results 4. Variance- is called the standard deviation squared; a
draw. 3. Wrong measure of variability.
4. incorrect patient transcription of the
identification patient’s data and TERMINOLOGIES:
5. Wrong specimen laboratory results.
container 1. Median – is the value of the observation that divides the
6. Incorrect observations into two groups, each containing equal
anticoagulant to numbers of observations. It is the midpoint of a distribution.

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 2. Mode – is the most frequent observation “Proper patient identification is the first step in sample collection”
3. Inferential statistics – are used to compare the means or this is the prime factor in order to attain accurate results in the
standard deviations of two groups of data. clinical laboratory.
4. t-test – is used to determine whether there is a statistically Patient Identification Procedures:
significant difference between the means of two groups of 1. Conscious Inpatients
data.  Verbally ask their full names.
5. f-test – is used to determine whether there is a statistically  Verify the name using the identification bracelet
significant difference between the standard deviations of which includes first and last names, hospital/ unit
two groups of data. number, room/bed number and physician’s name.
2. Sleeping patients
*Measures of center = (MMM)
 They are identified in the same manner as
*Measures of spread = range, standard deviation, coefficient of
conscious inpatients
variation
 They must be awakened before blood collection.
*Standard Deviation = 1SD: 68.3% 2SD: 95.4% 3SD: 99.7%
3. Unconscious, Mentally Incompetent Patients
*T and F tests = (TAM) (FPSD)
 They identified by asking the attending nurse or
Class of fire
relative; identification bracelet
CLASS OF FIRE MATERIAL EXTINGUISHER 4. Infants and children
A Ordinary water, dry chem.,  A nurse or relative may identify the patient, or by
combustible foam, loaded steam means of an identification bracelet.
materials (papers 5. Outpatient/ Ambulatory Patient
and woods)  Verbally ask their full names, address or birth date,
B Flammable organic Dry chem., CO2,
and countercheck with driver’s license or ID card
chemicals (organic loaded steam, halon
solvents) with photo
C Electrical Dry chem., CO2, halon  If the patient has identification card or bracelet,
D Combustible metals same manner as with hospitalized patients.
K Grease, oil and fats
*Type E = cannot be put off/ out → detonation (ARSENAL FIRE) General methods of blood collection:
REMEDY: let it burn out  Human body contains approximately 5 quarts (4.73
L) of whole blood
SPECIMEN COLLECTION AND HANDLING  Adult males: 5-6L of whole blood

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  Adult females: 4-5L of whole blood * Disinfection: circular motion
*Whole blood is composed of approximately: * Ethanol testing: benzalkonium chloride solution ( Zephiran
a. 60% of plasma – 3 quarts or 2.84L chloride, 1:750)
b. 40% of cells – 2 quarts or 1.89L * Needle insertion 15-30 degree angle
I. Arterial Puncture (Patient’s artery) *Apply pressure for 3-5 minutes at the site of venipuncture
 Arterial blood – oxygenated blood with a bright red color
 Blood sample is collected without tourniquet CARBOHYDRATES
 Preferred specimen for blood gas analysis and blood pH
measurement. Processes involved in carbohydrate metabolism:
 Sites: 1. Glycolysis
a. radial artery  Metabolism of glucose molecule to pyruvate or lactate to
b. brachial artery energy
c. femoral artery 2. Gluconeogenesis
d. scalp artery  Formation of glucose-6-phosphate from non carbohydrate
e. umbilical artery sources
* Arterial bleeding is the hardest to control and usually requires 3. Glycogenolysis
special attention.  Breakdown of glycogen to glucose for use as energy
II. Venipuncture (Patient’s vein. 4. Glycogenesis
 Venous blood – deoxygenated blood with a dark red color.  Conversion of glucose to glycogen for storage
 Sites: 5. Lipogenesis
a. antecubital fossa region  Conversion of carbohydrates to fatty acids
b. veins on the wrist and dorsal aspect of hands 6. Lipolysis
c. vein on the ankle (except for diabetic patients)  Breakdown of fats
* Median cubital vein – best site
Cephalic vein- second choice Hormones involved:
Basilic vein- third choice 1. Increases: glucagon, epinephrine, cortisol, growth hormone,
thyroxine
*Tourniquet application – 3-4 inches above the site, never leave the 2. Decreases: Insulin
tourniquet longer than one minute. 3. Regulator: Somatostatin
*Blood pressure cuff: 60mmHg

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* Insulin- Produced by the beta cells of pancreas Polyuria with high urine specific Polyuria with low urine specific
* Glucagon- Produced by the alpha cells of the pancreas gravity gravity
*Fasting blood Sugar: 8-10 hours of fasting
*Standard clinical specimen: Venous plasma glucose *Diabetes mellitus is associated with (PPP)
*at room temperature 7 mg/dL/ hr (0.4 mmol/L/hr) Types of Diabetes Mellitus:
*4 degrees Celsius: 2 mg/dL/hr Frequency 5-10% 90-95%
*HbA1c: monitoring glucose over the previous 2-3 months Risk factor Any (most common More common in
in children and advancing age
*Fructosamine: monitoring flucose over the previous 2-3 weeks
young adults)
Risk factor genetic, Genetic, obesity,
HYPOGLYCEMIA autoimmune, sedentary lifestyle,
 Decreased glucose levels environmental race/ ethnicity,
 Glycemic factors such as glucagon are released when the hypertension,
glucose levels reach 65-70 mg/dL dyslipidemia,
 Observable signs and symptoms of hypoglycemia appear polycystic ovarian
syndrome
when glucose levels reach 50-55 mg/dL
Pathogenesis Destruction of No autoimmunity,
 <50 mg/dL – diagnostic hypoglycemia value
pancreatic beta cells, insulin resistance
usually autoimmune and progressive
HYPERGLYCEMIA insulin deficiency
 Increased plasma glucose levels C- peptide levels very low or detectable
 Positive urine glucose undetectable
o Renal threshold limit for glucose is 140-160 mg/dL Pre- diabetes Autoantibodies Autoantibodies
(GAD65, IA2, IAA) absent
(160-180 mg/dL in some references)
may be present
= Increase Urine specific gravity
Medication therapy Insulin absolutely Oral agents, insulin
=Increase serum and urine osmolality necessary, multiple commonly needed
daily injections or
Diabetes mellitus vs. Diabetes Insipidus insulin pump
DIABETES MELLITUS DIABETES INSIPIDUS Therapy to prevent No known therapy Lifestyle (weight loss
Involves insulin Involves anti diuretic hormone or delay onset of and increased
Increased water excretion with Increased water excretion, no diabetes physical activity);
hyperglycemia hyperglycemia Oral medications can

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 be helpful  Can be found in the lungs as surfactant during
Previous names Insulin- dependent Non- Insulin respiration
DM dependent DM  Fatty acids
Juvenile- onset DM Adult- onset DM
 Building blocks of lipids
 Usually found in triglycerides, phospholipids and
LIPIDS
cholesterol
 Triglycerides (Triacylglycerol)
 Hydrophobic and water insoluble; serves as the
LIPOPROTEINS
main storage form of lipid in man(as found in
Normal lipoproteins:
adipose tissues)
 Chylomicron
 Do not have a any charge they are therefore known
 Largest and lightest among the lipoproteins; lowest
as neutral fats/ lipids
density (80-95% triglyceride be weight)
 Serves as: energy source, insulation or shock
 Transport exogenous triglycerides
absorber, as integral part of the cell membrane
 Causes non fasting lipemia ( turbidity of serum in
 Cholesterol
non fasting patients)
 Does not readily catabolized by most cells and
 No charge; remains in the origin during
therefore, does not serves as a source of fuel
electrophoresis
 Serves as: part of the cell membrane, fat absorption
 Very Low Density Lipoprotein
(fat emulsification), can be converted to steroid
 Pre- beta lipoprotein
hormones (glucocorticoids, mineralocorticoids and
 Transport endogenous triglycerides ( triglycerides
estrogens), can be transformed to vitamin D3 in the
produced by the body)
skin by irradiation from sunlight
 Low Density Lipoprotein
 AKA Bad cholesterol; Beta lipoprotein
*Cholesterol (cholesteryl) esters or esterified cholesterol: 60-70% of
 Transport cholesterol from the liver to the cells of
the total cholesterol in the body
the body
* Free cholesterol: 30-40%
 Directly proportional to the risk of atherosclerosis
 Phospholipids
and coronary heart disease ( higher LDL means
 Important constituent of cell membrane
higher risk)
(phospholipid bilayer), contains a polar and non
 High Density Lipoprotein
polar regions
 Smallest and the heaviest among the lipoproteins

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  Fastest towards the anode; alpha protein ; good
cholesterol
 Inversely proportional to the risk of atherosclerosis
and coronary heart disease ( Lower HDL means
higher risk)
 Responsible for reverse cholesterol transport;
transports cholesterol from the cells to the liver
Abnormal lipoproteins:
 Lp(A)
 Sinking prebeta lipoprotein
 Beta- VLDL
 Floating beta lipoprotein
 VLDL richer in cholesterol than in trigylcerides
 LpX
 Found in patients with obstructive biliary disease
*Ultracentrifugation: separates lipoproteins based on their
densities; reference method for lipoprotein analysis
*Standing Plasma Test: Used to detect chylomicrons or VLDL
*LDL = total cholesterol –HDL – (TG/5)
*Acceptable CV for lipid analysis: TC = 3%, LDL & HDL = 4%, TG = 5%
ENZYMES
 These are substances that catalyzes a given chemical
reaction
 The reaction they catalyze are frequently reversible which
means which means that they can synthesize and
decompose molecules
 They are complicated types of protein in terms of both
structure and function
 They easily denatured with varying molecular weight and
mass
 These enzymes are amphoteric which are capable of
ionizing either as acid or base
 They are synthesized in an inactive state and operates in the
presence of a cofactor
 Enzymes are found in all body tissues, they appear in the
serum following cellular injury or they may come from
degraded cells thus changes in enzyme concentration
reflects changes in state of health.
Definition of terms:
a. Coenzymes - non-protein organic biochemical that takes
part in the enzyme reaction.
 Essential to the catalytic activity as a CO-SUBSTRATE
 Diffusible, heat stable, low molecular weight that when
combined tightly to enzymes, the coenzyme will be called
Prosthetic group
E.g. NAD, Pyridoxal phosphate
b. Activators - Inorganic ionic cofactor
 increase the catalytic activity of an enzyme when it binds to
specific site
 Metabolic regulator of enzyme reaction
 Usually metal ions (esp. divalent cations)
E.g. Mg++, Na+, K+, Zn++
c. Holoenzyme - the combined enzyme & coenzyme
d. Apoenzyme - Enzyme without a cofactor
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 e. Prosthetic Group - A coenzyme that cannot be removed 2. First order reaction – reaction rate is directly proportional to
from its attachment to an enzyme using dialysis substrate concentration.
f. Substrate - Substance acted upon by an enzyme & is
converted into a new substance I. PHOSPHATASES
g. Product - Substance derived from a transformed substrate A. Alkaline Phosphate/ Alkaline Orthophosphoric Monoester
h. Active site - Site where substrate interacts with enzymes Phosphohydrolase
i. Allosteric site – Site other than the active site that may lead  Major Tissue source: liver, bone, placenta, intestinal, renal
to either attachment of substrate to the enzyme’s active  Diagnostic significance:
site or inhibition of attachment.  When total ALP levels are increased, it is the major
j. Isoenzymes – different form of an enzyme with different liver fraction that is most frequently elevated =
genetic origins but catalyze the same reaction. obstructive jaundice.
k. Isoforms – Results when an enzyme is subject to different  For bone disorders, highest elevations occur in
post-transitional modification Paget’s disease (osteitis deformans)
CLASSIFICATION OF ENZYMES  Isoenzymes:
1. Oxidoreductase (dehydrogenases/ Oxidases) 1. Liver ALP – most anodal
2. Transferase ( kinase/ transaminase) 2. Bone ALP
3. Hydrolase 3. Placental ALP
4. Lyase ( decarboxylase) 4. Intestinal ALP – least anodal
5. Isomerase B. Acid Phosphatase/ Acid Orthophosphoric Monoester
6. Ligase Phoshohydrolase
 Useful in forensic clinical chemistry, in the investigation of
Enzyme theories: rape cases – vaginal washings are examined for seminal
1. Emil Fisher’s/ Lock and key Theory – The sape of the key fluid acid phosphatase (ACP) activity, which can persist for
(substrate) must fir into the lock (enzyme) up to 4 days
2. Kochland’s/ Induced Fit Theory – Based on the substrate binding  Diagnostic significance:
to the active site of the enzyme  Prostatic carcinoma, forensic chemistry
II. TRANSAMINASES/ TRANFERASES
Enzymatic reactions: A. Aspartate Aminotransferase (AST)
1. Zero order reaction – reaction rate depends only on enzyme  Formerly known as Serum glutamic-oxaloacetic
concentration. transaminase (SGOT)

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  Often used in conjunction with ALT for hepatocellular
disorders
B. Alanine Aminotransferase (ALT)
 Formerly known as Serum glutamic-pyruvic transaminase
(SGPT)
 Specific for liver disease compared to AST
 The highest concentration is in the liver
*De Ritis ratio (ALT: AST) is >1.0 = Acute hepatitis
III. AMYLASE/ Alpha-1-4 Glucan-4-Glucohydrolase (AMS)
 It catalyzes the breakdown of starch and glycogen
 Smallest enzyme in size
 Earliest pancreatic marker
 Isoenzymes:
 S-type (ptyalin) and P-type (amylopsin)
 Major tissue sources: acinar cells of the pancreas and the
salivary glands
*Substrate: starch
IV. LIPASE( LPS) / Triacylglycerol acylhydrolase
 Most specific pancreatic marker- secreted exclusively in the
pancreas; not affected by renal disorders.
V. LACTATE DEHYDROGENASE (LDH)
 Electrophoresis:
LD5 (origin) – LD4 – LD3 – LD2 – LD1 (Anode)
 Concentration in serum: LD2>1>3>4>5
*LD flip (LD1>LD2) is associated with AMI
* Wacker – forward reaction
*Wrobleuski La Due – Reverse reaction
VI. CREATINE KINASE/ Creatine phosphokinase
 Highest elevation of total CK is seen in Duchenne’s muscular
dystrophy
 Isoenzymes: CK-BB (brain type) , CK-MB (hybrid type), CK-
MM (muscle type)
*Tanzer-Gilvarg – forward reaction ( Pyruvate kinase , LDH)
*Oliver- Rosalski – reverse reaction (Hexokinase, G6PD)
VII. GAMMA GLUTAMYL TRANSFERASE
 Sensitive indicator of alcoholism (most sensitive marker of
acute alcoholic hepatitis.
VIII. Cholinesterase (CHS)/ Pseudocholinesterase
 Marker for insecticide/ pesticide poisoning
IX. GLUCOSE-6- PHOSPHATE DEHYDROGENASE (G-6-PD)
 It is an newborn screening marker.
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