CLINICAL MICROSCOPY

A field of Clinical Pathology that deals with the study of all

body fluids besides blood. Clinical Microscopy includes:
Urine analysis
CSF analysis
Seminal Fluid analysis
Serous fluid analysis (transudates & exudates)
Synovial fluid analysis
Amniotic fluid analysis
Gastric Fluid analysis
Sweat analysis
Fecal analysis
Sputum & bronchial lavage analysis and Others
Objectives
Formation/Physiology of each body fluid
Sampling and collection of each body fluid
Normal characteristics/features of each body fluid
Composition/constituents of each body fluid
Laboratory Determinations done on each body fluid
Clinical significance of these tests
Safety procedures and quality controls in CM
URINE
RENAL PHYSIOLOGY
Nephrons
= functional unit
= 1-1.5 M/kidney
Renal Functions:
1. Renal blood flow
2. Glomerular filtration
3. Tubular reabsorption
a. Active transport:
PCT: glu, amino acids, salts
ALH: Chloride
DCT: Sodium
b. Passive transport
H2O: throughout the nephron except ALH
Urea: PCT and ALH
ALH: Sodium

4. Tubular secretion
- Result: Blood is cleared from waste products;
water, acid
base balance and electrolyte
balance are maintained
Other functions:
- erythropoiesis
- BP maintenance
- Calcium level maintenance
Formation & Physiology
= ultrafiltrate of plasma
= rate:
1,200 1,500 ml/min of urine from a renal
BF of 170,000
ml/day (20-25% total blood volume)
= factors affecting composition: diet, activity, body
metabolism,
endocrine function
Composition:
1. Urea, creatinine, uric acid = use to ID a fluid as urine
2. Inorganic subs (Na, Cl, K)
3. Water
Major organic component: urea
Major inorganic component: Cl4. Albumin (150mg/dL)
Urobilinogen (1 mg/dL or 1 Ehrlich unit)
5. Formed elements
few squamous, transitional,renalepithelial cells/ lpo
few mucus/lpo
amorphous urate and phosphate crystals
0-2 red blood cells/hpo
0-5 white blood cells/hpo
0-2 hyaline casts/hpo
Sampling and Handling
Collection:
= volume: > 12 ml
= viability: < 2 hrs
Specimen Preservation:
1. Refrigeration
2. Chemical preservatives
= bactericidal, inhibit urease, preserve formed
elements
= NO best preservative; depends on the test
requested
Types of Urine Specimen
1. Random specimen = routine

2. First morning specimen = pregnancy test, orthostatic

proteinuria
3. Fasting specimen/ 2nd Morning = DM monitoring
4. 2 hr Postprandial Specimen = DM monitoring
5. Glucose Tolerance specimen = with OGTT
6. 24 hr (Timed) specimen = quantification; chemical tests
7. Catheterized urine = Bacteriologic study
8. Midstream clean catch urine = Bacteriologic study
9. Suprapubic aspiration = Bacteriologic study (anaerobes)
10.Three glass collection = prostate cancer
11.Pediatric specimen
12.Drug specimen collection
2 Main Types of Urinalysis
1. Dipstick urinalysis - Screening test; point of care
2. Routine Urinalysis (Wet urinalysis)
-Confirmatory test: performed in laboratories
-Chemistry + Microscopy
a. Manual
b. Automated
-semi-automated
-Fully automated:
Ex: IRIS work stations light scattered,
fluorescence &
impedance
UF 100 Sysmex - Flowcytometry
Other Urine Laboratory Tests
Urine Microbiology
Urine cytology
Cytodiagnostic urinalysis
Image cytometry and DNA analysis
Flow cytometry
ROUTINE URINALYSIS
I- Physicochemical examination
- clarity, color, specific gravity, odor
- glu, proteins, pH, etc
II- Microscopic Study
- cells, crystals, casts, microbes
PHYSICAL EXAMINATION
Urine volume
Normal urine volume: 600-2,000 ml/day
night urine: < 400 ml

Factors affecting urine volume:

1. state of hydration
2. ADH
3. Increase amounts of dissolved solids (glu & salts)
Clinical significance of Urine Volume:
ANURIA:
= near complete cessation of urine formation
OLIGURIA:
= decrease in normal daily urine volume
= < 30 ml/hr; < 500 ml/day
Causes:
1. Pre-renal
-decrease renal BF
-DHN: vomiting, diarrhea, excessive sweating, severe
burns, edema
2. Renal
- GN, ATN, chronic renal failure
3. Post-renal
-obstruction: stones, BPH, tumors, severe DHN
POLYURIA:
= increase in normal daily urine volume
= >2000 ml
= DM, DI, diuretics, caffeine, alcohol
NOCTURIA:
= increase in nocturnal urine output
= > 500 ml at night; sp. gr < 1.018
= DM
Mechanisms for increase urine output:
defective hormonal regulation of volume homeostasis
Defective renal salt/water absorption
Osmotic diuresis
Color
Normal: straw to amber
Indicator of individuals state of hydration
Normal urine pigments:
1. urochrome (yellow) = major
-increased: fever, starvation, thyrotoxicosis
2. uroerythrin (pink)
3. urobilin (orange-red)
Abnormal urine pigments: Strasinger

Notes on Urine color

Red urine - In porphyria varies
Dark-brown or cola-colored
-Rhabdomyolysis
-L-DOPA medication, alkapton, melanin
Clarity or Appearance
Normal: clear
Non-pathologic causes of turbid urine:
o squamous epith. cells
o mucus threads
o amorphous PO4 and urates
o sperm cells in females
o fecal contamination
o radiographic contrast medium
o talcum powder/vaginal creams
Pathologic causes of turbid urine:
o rbc
o wbc
o bacteria
o yeast
o nonsquamous epithelial cells
o abnormal crystals
o lymph fluid
o lipids
o malignant cells
How to differ causes of urine cloudiness
Acidification:
Phosphates:
dissolves
Wbc:
persists
Bacteria:
persists
Filtration

Wbc:
resolved
Bacteria:
persists
Ether or chlorform
Chyluria:
resolved

Specific Gravity & Osmolality

SPECIFIC GRAVITY = measure relative proportion of
dissolved particles to total volume of specimen; density
Normal: 1.023 (1.003 -1.035)
Clinical significance: reflects the ability of the kidney to
concentrate and dilute urine
Isosthenuria:
1.010; severe renal damage
Hyposthenuria:
<1.007
Hypersthenuria:
>1.010
Methodology:
1. Urinometer = density
2. Refractometer = light wave scattered (refractive
index); <0.002
of urinometer rdng
3. Harmonic oscillation densitometry = frequency
of sound wave
4. Reagent strip = pKa changes of polyelectrolyte
5. Falling drop method (automated) = amt of time
it takes a drop
of urine to fall a fixed distance; direct
mtd; > accurate and precise
Osmolality
OSMOLALITY = measure number of solute particles per unit
of solvent; measures Na and Cl
NV: 500-850 mOsm/day
Methods: Freezing point depression method (most common)
Odor
Normal: aromatic (volatile acids)
Abnormal:
o Foul, ammonia-like
UTI, old specimen
o fruity, sweet
ketones (DM,
starvation,vomiting)
o maple syrup
maple syrup syndrome
o mousy
Phenylketonuria
o rancid
Tyrosyluria
o sweaty foot
Isovaleric acidemia

o
o
o
o

Cabbage/hops
rotting fish
Bleach
lack of odor

Methionine malabsorption
Trimethylaminuria
semen contamination
acute tubular necrosis (ATN)

Chemical Examination
1. pH
Normal: 4.5 8.0
Clinical significance: a measure of the kidneys ability to
maintain normal H ion conc in plasma and ECF
A. acid-base disorders
B. renal damage
C. renal calculi formation
D. treatment of UTI
E. precipitation and identification of crystals
F. determination of unsatisfactory urine specimen (pH
>9.0)
Causes of Acid and Alkaline urine
Acid urine
Alkaline urine
COPD
Hyperventilation
DM
Vomiting
Starvation
Renal tubular acidosis
Dehydration
Urease- producing
bacteria (Proteus)
Acid-producing bacteria
Vegetarian and fruit diet
(E. coli)
High protein diet
Old specimens
Cranberry juice
pH: Methods of detn
Reagent strip
pH electrode - More specific and sensitive
Titrable acidity of urine
2. PROTEINS
Normal: 2 - 10 mg/dL or up to 150 mg/24 hr
mainly albumin
Clinical Significance:
= has low max tubular rate of reabsorption (Tm)
= increase filtration of protein quickly saturates tubular
reabsorption
A. Pre-renal Proteinuria

- intravascular hemolysis (Hgb)

- muscle injury (myoglobin)
- severe infection & inflammation (acute phase
reactant proteins)
- multiple myeloma (BJP)
B. Renal Proteinuria
a. Glomerular disorder
- immune complex disorder (SLE, PSGN)
- amyloidosis
- toxic agents
- Nephrotic syndrome (> 4g/day)
- diabetic nephropathy
- renal hypertension- dhn, strenous exercise,
eclampsia, orthostatic
b. Tubular disorders
- Fanconis syndrome
- toxic agents/ metals
- severe viral infections
C. Postrenal Proteinuria
- lower UTI
- injury/trauma
-menstrual contamination
- seminal fluid contamination
-vaginal secretions
Protein Quantification
Microalbuminuria:
AER = 20-200ug/min or 20-200 mg/L
alb: crea ratio > 3.4 mg/mmol
4- 6x increased risk of CVD mortality in DM
Independent risk factor for renal mortality
Mild proteinuria = < 1 g/day
- Tubular damage in origin
Moderate proteinuria = 1 4 g/day
Heavy proteinuria = > 4 g/day
-glomerular damage in origin
Methods of detn
Reagent strips
Precipitation tests & Turbidimetric mtd
Dye binding methods
ICT
Electrophoresis


Methodology: Reagent strips vs Precipitation Tests for
Proteins
More sensitive but less specific than reagent strips
Can detect other proteins besides albumin (globulins,
BJP, glycoproteins, radiographic dyes)
Reagents: 5% HAc, 3% SSA or TCA
Manner of reporting:
(-) = no turbidity
< 5 mg/dL
(+/-)= trace
20 mg/dL
(1+) = faint turbidity only
50
mg/dL
(2+) = turbidity with granulation
200
mg/dL
(3+) = turbidity, granulation, flocculation
500
mg/dL
(4+) = clumps of precipitation
> 1.0
g/dL
3. GLUCOSE
Normal Renal Threshold: 180-200 mg/dL
Normal: minute amounts
Clinical Significance:
A. Hyperglycemia associated
a. decreased insulin
- DM
- Pancreatitis, pancreatic cancer
b. inhibition insulin
- acromegaly (increase growth hormone)
- Cushings syndrome (increase cortisol)
- Hypertyhroidism (increase T3 T4)
- Pheochromocytoma ( increase
- stress (CNS damage, MI)
- gestational diabetes (increase HPL)
B. Renal associated
- Fanconis syndrome
- Advanced renal disease
- Osteomalacia
- Pregnancy
Methodology: Reagent strip vs Colorimetric test

Note: Positive Clinitest and Negative Dipstick test is seen in

the following conditions:
1. non-glucose mellituria = inherited enzyme
deficiencies
2. Lactosuria = late pregnancy; lactation
3. Pentosuria = ingestion of certain fruits
4. Fructosuria = parenteral feedings with fructose
5. Other non-sugar reducing substances like ascorbic
acid
NB: sucrose = non-reducing sugar so does not react
with Clinitest
or dipstick test
4. KETONES
Normal: none to minute
Three forms:
a. acetoacetic acid (20%)
b. B-hydroxybutyric acid (78%)
c. acetone (2%)
Clinical significance:
A. Diabetic ketoacidosis
B. Insulin dosage monitoring
C. Starvation
D. Malabsorption/pancreatic disorders
E. Strenuous exercise
F. Vomiting
G. Inborn errors of amino acid metabolism
Methods of detn
Gerhardts test- Ferric chloride reaction
Reagent strips - Nitroprusside reaction in strips
Rotheras test - Nitroprusside reaction in test tubes
Acetest - Nitroprusside reaction in tablet form
NB: ALL methods do not detect B-hydroxybutyrates
5. BLOOD
Normal: rbc= 0-5/ hpf
Forms:
intact rbc or hemoglobin
dysmorphic rbc: glomerular dse
Urine starts to be pink: Hgb > 50 mg/dL
Hemosiderin pigments:
2-3 days after acute
hemolytic episode
Clinical Significance

A. Hematuria
- renal calculi
- GN, pyelonephritis
- trauma & tumors
- toxic chemicals
- anticoagulants
- strenuous exercise
B. Hemoglobinuria
- transfusion reactions
- hemolytic anemias
- severe burns
- infections/malaria
- strenous exercise
C. Myoglobinuria
- muscular damage
- prolonged coma
- muscle wasting disease
- convulsions
- alcohol overdose
- drug abuse
- strenuous activities
NB: massive hemoglobinuria & myoglobinuria is toxic to the
kidneys, can cause Acute renal failure (ARF)
6. BILIRUBIN
Normal: conjugated form (B2)= <0.5 mg/dL
Clinical significance:
A. Pre-hepatic jaundice
a. Hemolytic anemia
b. severe muscle destruction
B. Hepatic jaundice
a. Hepatitis
b. Cirrhosis
c. Other liver diseases/disorders
C. Post-hepatic or obstructive jaundice
a. All stones
b. Malignancy
7. UROBILINOGEN
Normal: 1 mg/dL
Clinical significance:
A. Early detection of liver disease

B. Liver disorders, hepatitis, cirrhosis, malignancies

C. Hemolytic disorders

Urine bilirubin vs urobilinogen in jaundice

Type of jaundice
Urine
Urobilinoge
bilirubin
n
Pre-hepatic(haemolytic
negative
+++
dse)
Hepatic (liver damage)
+ or ++
Post-hepatic (bile
+++
Normal to
duct obstruction
decreased
8. NITRITE
Normal: colony count = < 100,000/ml
NO2= < 0.05 mg/dL
Clinical significance:
A. UTI
B. Evaluation of antibiotic treatment
C. Monitoring for patients at high risk for UTI
D. Screening urine specimen for culture
9. LEUKOCYTE ESTERASE
Normal: wbc =0-2 or 0-5/hpf
Clinical significance:
A. UTI (bacterial or non-bacterial)
B. Inflammation of the urinary tract without bacteruria
C. Screening specimen for culture
MICROSCOPIC STUDY
Methods:
1. Microscopy
a. bright field microscopy - routine urinalysis
-Sampling: 0.02 ml reconstituted
-scan 10 fields/lpf or hpf; stained or unstained
b. Phase contrast microscopy
- casts, mucus threads, Trichomonas
c. polarizing microscopy
- cholesterol in crystals and casts
d. Interference-contrast microscopy

- three dimension image for crystals

e. Cytology detection of malignancies
2. Automated Commercial Systems
Microscopic Sediment stains (Supravital stains)
1. Sternheimer-Malbin stain = crystal violet and safranin
O
= casts and cells
2. 0.5% toluidine blue = various cell component
= differentiates wbc vs renal cells
3. Sudan III or oil red O = neutral lipids, oval fat bodies,
TG
4. Prussian blue = hemosiderin (Fe)
5. Hansel stain = methylene blue, eosin-Y in methanol
= eosinophils
URINARY SEDIMENTS
1. Cells: wbc, rbc, epithelial cells, bacteria, sperm cells
2. Casts: Tamm-Horsfall protein matrix, formed at the
distal convoluted tubules
3. Crystals
4. Miscellaneous: mucus threads, fat droplets
5. Epithelial cells

Pathologic cells in urine

Transitional cells
Renal cells

RBCs & WBCs (pus cells)

URINARY CRYSTALS
Pathologic Crystals (Acidic urine)
-cholesterol, cysteine, tyrosine, leucine
Non-pathologic Crystals (Acidic urine)
-Calcium oxalate
-Uric acid
Non-pathologic crystals (Alkaline urine)
-Triple phosphate
-Calcium phosphate
-Ammonium biurate
-Calcium carbonate
URINARY CASTS
Factors which favor protein cast formation:
low flow rate
high salt concentration
low pH
NB: all of which favor protein denaturation and
precipitation,
particularly that of the TammHorsfall protein.
CYLINDROIDS: Protein casts with long, thin tails formed
at the
junction of Henle's loop and the distal
convoluted tubule
Urinary Casts
Hyaline cast
Wbc cast
Rbc cast
Coarse granular cast
Fine granular cast
Broad cast
Broad waxy cast
Fatty cast
Manner of Reporting
How to quantify:
Numerical
Qualitative:
o Occasional
o Few
o Some

o Many
o TNTC
Epithelial cells, mucus threads, bacteria,
crystals, yeast cells
Objective of microscope to use:
Low power objective
o Epithelial cells
o Mucus threads
o Crystals
o Casts
High power objective
o Rbc
o Wbc/pus cells
o Yeast cells
o bacteria
Reference values
rbc:
0-2/hpo
wbc/pus cells:
0-5/hpo
hyaline cast:
0-2/hpo
bacteria:
occasional-few/hpo
crystals:
neg/lpo
Urinary casts
- Telescoped urinary sediment: is one in which red cells,
white cells, oval fat bodies, and all types of casts are
found in more or less equal profusion.
- Associated with:
lupus nephritis
malignant hypertension
diabetic glomerulosclerosis
rapidly progressive glomerulonephritis
RENAL FUNCTION TESTS
First consideration in kidney damage
Do URINALYSIS first!
Advantage:
-tells us what is wrong
-pinpoints site of defect
-its cheap
Disadvantage:
-doesnt tell us whether the damage is

-primary or secondary renal disease

-localized or generalized
Do RENAL FUNCTION TESTS to resolve the said
disadvantages
A. Glomerular Filtration Tests
CLEARANCE TESTS = volume of plasma from which a
measured amount of subs can be completely eliminated
into the urine per unit time
Normal urinary clearance: 120ml/min
Features of a good CT:
a. subs is neither reabsorbed or secreted by the
kidneys
b. consistency of its plasma level
c. sub availability in the body
d. availability of the test
Specimen:
o 24 hour urine: excretion of crea/urea is not
consistent
o Alternative: 2 hr fasting urine/catheterized urine
Collection and handling:
o 1st urine discard; collect all succeeding urine; include
last voided urine at the end of the 24 hr time
o refrigerate
1. Urea Clearance Test
- Earliest
- Disadv:
40% reabsorbed back; Affected by diet,
diurnal changes
2. Inulin Clearance Test
- Polymer of fructose
- Adv: stable
- Disadv: Exogenou, Availability
3. Radionuclide Clearance test
- Adv: no need for 24 hr urine
visualize glomerular function
- Disadv: exogenous, expensive
4. B2- macroglobulin Clearance Test
- Adv: more sensitive and specific
- Disadv: expensive, availability
not reliable for patients with
immunologic problems and malignancies
5. Creatinine Clearance test (CCT)

routinely used by lab

- adv: Endogenous, stable, inexpensive
- disadv:
secreted by the tubules (20-30%)
chromogen = false increase
bacterial disintegration
meat diet increase result
not reliable in muscle wasting d/o
- Mtd: Jaffes alkaline picrate test
- NV:
male
107-139 ml/min
female
87-107 ml/min
- Calculation:
C= UV
P
where:
C = clearance/min
U = urine crea conc
P = plasma crea conc
V = vol/min = vol of 24 hr urine/1440
Clinical Significance of CCT:
1. determine EXTENT of glomerular damage
mild:
60-80% of NV
moderate: 40-60% of NV
severe:
20-40% of NV
2. monitor effectiveness of treatment
3. determine feasibility of administering treatment
Application of CCT:
- Measures overall renal impairment but will reflect a
predominant GLOMERULAR DAMAGE!
- Shows EARLY lesions of the kidney without increase in
BUN level
- Use to follow up the clinical course of the disease
B. Tubular Reabsorption Tests
- first fxn affected by renal disease and last fxn to be
lost
- important in differentiating prerenal azotemia from
renal failure
CONCENTRATION TESTS = test to determine ability of
the tubules to reabsorb
essential salts and water
that have been filtered out by
the glomerulus
= urine is plasma ultrafiltrate
= sp. gravity of urine:

after GF = 1.010
after TR & TS = higher; its concentrated
Concentration Tests
1. Fishberg Concentration Test
= measures sp. Gr. After 24hr water deprivation
2. Mosenthal Concentration Test
= compares vol & SG of day & night urine samples
3. Free Water Clearance Test
= measures osmolar clearance
= determines ability of tubules to respond to state of
hydration
= Interpretation of results:
-2
= dehydration
0
= no renal concentration/ dilution
+2
= over hydration
4. Specific Gravity
= simple, readily available, inexpensive
= NV: 1.025 (1.025- 1.035)
=disadv: does not measure dissolved solutes
= Clin significance:
a. initial evaluation of early renal disease
b. monitoring course of the disease
c. monitoring electrolyte and fluid tx
d. isosthenuria (1.010):
- Diabetes insipidus
- diuretic phase of ATN
- hyperthyroidism
- sickle cell anemia
- salt- restricted diets
5. Osmolality
= best quantitative measurement of renal
concentrating ability
= measures also urea, Na, Cl which contributes to
urine conc.
= measured by Osmometers
a. Freezing Point Osmometer
b. Vapor pressure osmometer
= NV: 800-1,300 mosm/L
Normal serum:urine osm = 1:1 to 3:1
C. Tubular Secretion Tests

subs must be completely removed from the blood by

the peritubular capillaries
1. Para-Aminohippuric Acid Test (PAH)
= derived from PSP test
= adv: non-toxic
do not bind strongly to plasma proteins
= NV: 600-700 ml/min
2. Titrable Acidity & Urinary NH3
= measures PCT (H+ secretion) and DCT (NH4+
secretion) functions
= NV: 70 meq/day
= Clin. Significance:
Metabolic acidosis= inability to produce an acid
urine
INTRAVENOUS PYELOGRAPHY (IVP):
- An x-ray study where a dye is injected IV to visualize
the urinary system
- Dye is secreted by the tubules. If it is not secreted =
renal tubular damage
- NB: DO NOT use in patients with uremia
- Use to diagnose:
o Obstruction
o Masses/malignancies
o Sizes of kidney (atrophy/hydronephrosis)
o Tubular secretion capacity of the renal tubules
TESTS for SEVERE GLOMERULAR or TUBULAR DAMAGE
or BOTH
1. BLOOD UREA NITROGEN (BUN) det2.
2. BUN CREATININE detn
AZOTEMIA = Increase BUN and creatinine level in the blood
and urine
UREMIA =Azotemia + clinical S/Sx of renal failure
BLOOD UREA NITROGEN
= First to increase in early renal damage
=Not reliable if tested alone because it is affected by diet
and other factors
CREATININE
= Tends to increase later in severe renal damage
=If elevated may suggest chronicity
=Factors affecting BUN do not affect crea
Commonly requested alone

AZOTEMIA 3 Classifications:
a. PRE-RENAL AZOTEMIA
- Problems before the kidney
- Associated with
- Decrease blood volume and renal flow
- Increase protein intake or catabolism
- Seen in: Traumatic shock, electric shock, severe DHN,
acute cardiac decompensation, overwhelming
infection, Chemotherapy, GIT hemorrhage
b. RENAL AZOTEMIA
- Direct involvement of the kidney
- Chronic, diffuse, bilateral
- Seen in: Chronic Glomerulonephritis, Bilateral chronic
pyelonephritis, ATN, Severe acute GN
c. POST RENAL AZOTEMIA
- Obstruction anywhere from the ureter to pelvis
- Seen in: Calculi, Tumors (Primary to lower GUT BPH or
prostatic Ca)
d. TERMINAL AZOTEMIA
- Azotemia seen in patients with terminal cancer
- BUN is increased to a uremic level
- May be due to increased protein catabolism

INBORN ERROR OF METABOLISM

(NEWBORN SCREENING)
URINE SCREENING FOR METABOLIC DISORDERS
2 Pathways:
1. Overflow disorders disruption of metabolic pathway--increase plasma conc of unmetabolized subs--increase conc in urine
ex: inborn error of metabolism
2. Renal disorders malfunctions in tubular reabsorption
OVERFLOW METABOLIC DISORDERS
A. Amino acid disorders
1. Phenylalanine-tyrosine disorders
a. Phenylketonuria
b. Tyroslyuria
c. Alkaptonuria
d. Melanuria
2. Branched chain Amino Acid disorders
a. Maple syrup urine disease

b. Organic acidemias
3. Tryptophan disorders
a. Indicanuria
b. 5-Hydroxyindoleacetic aciduria
4. Cystine disorders
a. cystinuria / cystinosis
b. Homocystinuria
5. Porphyrin disorders
B. Mucopolysaccharide disorders
a. Sanfilippos syndrome
b. Hurlers syndrome
c. Hunters syndrome
C. Purine disorders
a. Lesch-Nyhan disease
D. Carbohydrate Disorders
a. Pentosuria
b. Galactosuria
c. Lactosuria
d. Fructosuria
SAMPLING
Routine Newborn Screening test
- typically performed at least 12 hours and generally 2428 hours after birth
- Repeat: 2 wks old
Sample: Urine; sweat; blood
PHENYLALANINE METABOLISM DISORDERS
PKU
Tyrosluria
Melanuria
Alkaptonuria
A. PHENYLKETONURIA (PKU)
- an autosomal recessive genetic disorder
- Incidence:
1 in 15,000 births
1 in 4,500 births (Ireland)
1 in 100,000 births (Finland)
- Etiology: deficiency in phenylalanine hydroxylase (PAH)
in the liver enzyme is necessary to metabolize the amino
acid phenylalanine to the amino acid tyrosine.

if deficient, phenylalanine accumulates and is converted

into phenylpyruvate (also known as phenylketone),
phenylacetate, and phenylethylamine which is detected
in the urine.
- PAH is found on chromosome number 12.
- Manifestations:
o Brown, mousy odored urine (phenylacetate)
o Intolerance to milk
o Microcephaly, mental retardation, seizures, learning
disabilities
o albinism, eczema
o Damage done is irreversible so early detection is
crucial.
- Treatment: diet low in phenylalanine and high in tyrosine
can be a very effective treatment. There is no cure.
- Tests:
o Guthries test: (+) growth
o FeCl3 strip test (Phenistix): blue to green color
o High-performance liquid chromatography (HPLC)
B. TYROSYLURIA
- Etiology: tyrosine transaminase deficiency - Increased
tyrosine and its by products (para-hydroxyphenyl
compounds)
- Manifestations:
o Rancid urine
o Mental retardation
o Liver disease
o Tyrosyluria
- Tests:
o Nitrosonapthol test: red color
o (+) tyrosine crystals in urine
o HPLC
C. Melanuria
- Etiology: tyrosine transaminase deficiency
Excess tyrosine --- melanin
- Manifestations:
o Black urine
o Hyperpigmentation
o Mental retardation
- Tests:
o Fe Cl3 test: blue color with black precipitate

o Na nitroprusside: red color

D. Alkaptonuria
- Etiology: homogentisic acid oxidase deficiency --increased homogentisic acid (alkaptons)
- Incidence: more common in Slovakia and the Dominican
Republic
- Manifestations:
o Black urine
o Mental retardation
o damage to cartilage (ochronosis, leading to
osteoarthritis) and heart valves
o kidney stones
- Tests:
o FeCl3 test: transient blue color
o AgNO3 test:black color
o HPLC
ORGANIC ACIDEMIAS
Maple Syrup Urine Disorder
Organic Acidemias
A. Maple Syrup Urine Disorder (MSUD) or Branched
Chain Amino Acid Disorder
- Etiology: deficiency of branched-chain -keto acid
dehydrogenase (BCKDH)
- increased branched-chain amino acids (leucine,
isoleucine, and valine) and their toxic by-products in the
blood and urine
- Incidence: 1 in 80,000 birth
- Manifestations:
o Maple syrup odor urine
o Liver disease
o Metabolic encephalopathy (poor feeding, vomiting,
dehydration, lethargy, hypotonia, seizures,
ketoacidosis, and neurological decline)
- Tests:
o (+) leucine crystals
o 2-4 DNPH test:
yellow precipitate
B. Organic Acidemias
- Etiology: oxidative decarboxylase deficiency
-Increased organic amino acids (isovaleric acid, propionic
acid, methylmalonic acid)

Manifestations:
o Sweaty odor
o Vomiting, hypoglycemia, metabolic acidosis, ketonuria,
uremia
Tests:
o P-nitroaniline test:emerald green

TRYPTOPHAN DISORDERS
Indicanuria
5-Hydroxyl indole acetic acidemia (serotonin)
A. Indicanuria (Blue diaper baby syndrome)
- Etiology: autosomal or X-linked recessive trait; genetic
metabolic disorder characterized by the incomplete
intestinal breakdown of tryptophan
-intestinal obstruction
-Malabsorption
-Hartnups disease
- Manifestations:
o Digestive disturbances like vomiting, diarrhea
o irritability and visual difficulties
o kidney disease
o bluish urine-stained diapers
- Tests:
o Exposure to air:
indigo blue
o FeCl3: deep blue or purple
B. 5-Hydroxy Indole Acetic Acidemia (Serotonin)
- Etiology: Malignancy of the argentaffin cells --- increased
5-HT
- Manifestations:
o tachycardia [fast heart rate]
o a rise in blood pressure,
o bronchospasm going as far as asthma attack,
o increased intestinal peristalsis (contractions and
dilations of the intestines to move the contents
onwards)
o increased sensitivity to pain
o increased aggression
- Test:
o 2,4-DNPH Test:
purple to black color
CYSTINE DISORDERS

A. Cystinuria and Homocystinuria

- Etiology: defective tubular reabsorption
Increased arginine, lysine, ornithine
- Manifestations:
o Liver disease
o Calculi formation
- Tests:
o (+) cystine crystals and stones in urine
o Cyanide-nitroprusside test: red to purple
PORPHYRIN DISORDERS
- Etiology: Defect in heme synthesis due to:
a. Inherited enzyme deficiency
b. Lead poisoning (Increased in uroporphyrin,
coproporphyrin, protoporphyrin)
- Manifestations: red urine varies
o Port-wine: Congenital erythropoeitic porphyria and
porphyria cutaneous tarda
o Acute intermittent hepatic porhyria: normal --- darkens
on standing
o Normal: Lead porphyrinuria anemia
- Tests:
o Ehrlichs test: cherry red
o Fluorescence test under UV: pink/violet/red
fluorescence
o To differentiate porphobilinogen from hemoglobin:
Hoeschs test and Watson-Schwartz test
CALCIUM DISORDER
- Etiology: endocrine problems: Hyperparathyroidism,
Osteoporosis, Multiple myeloma, Bone malignancies
- Manifestations:
o Muscle twitching
o Arrythmia
o Pathologic fractures
- Tests:
o Sulkowitch test:
white precipitate
MUCOPOLYSACCHARIDE DISORDERS
Hurlers syndrome
Hunters syndrome

San Filippo syndrome

Etiology: enzyme deficiencies
Increased dermatan sulfate, keratan sulfate,
heparan sulfate
- Manifestations:
o Hurlers syndrome: short stature, weakness & paralysis
o Hunters syndrome: muscle weakness, corneal
deposits, loss of vision
o San Filippos syndrome: severe mental retardation
- Tests:
o Acid albumin test: turbidity
o Cetyltrimethylammonium bromide test (CTAB):
turbidity
o Metachromatic staining spot test: blue spot
-

PURINE DISORDERS
A. GOUT
- Etiology: purinase deficiency --- increased uric acid
- Manifestations:
o Orange sand pebbles in diapers
o Motor defects
o Gout
o Renal calculi formation
o Mental retardation
- Uric acid
- Tests:
o (+) uric acid crystals in urine
o Increased BUA
CARBOHYDRATE METABOLISM DISORDERS
A. Hydroxylases Deficiency Disorders
- Etiology: hydroxylase deficiency
Ex: lactase, maltase
- Manifestations:
o Malabsorption
o Non-glucose mellituria
o Failure to thrive
o Mental retardation
o Liver disease
o cataract
- Tests:

o
o
o

Non-glucose reduction tests:

Benedicts test
Clinitest

PREGNANCY TEST
based on the fact that the placenta (trophoblastic cells)
secrete Human Chorionic Gonadotropin (hCG)
- a hormone that has a luteinizing action on the ovarian
follicles
- Can occur in 3 subunits:
B-hCG subunit
Alpha-hCG subunit
Whole hCG molecule
Indications
a. Early diagnosis of pregnancy (1st trimester)
b. Ectopic pregnancy
c. Evaluation of threatened abortion and after evacuation
of incomplete abortion
d. Guidance for the diagnosis and treatment of
trophoblastic tumors
e. Evaluation of selected non-trophoblastic tumors
Methods of Pregnancy test
a. Bioassay
b. Immunoassay
- Agglutination Immunoassay
- Radioimmunoassay
- ELISA
c. Radioreceptor Assay
1. BIOASSAY
- In vivo test; done in reference lab only
- Limitations: very crude, expensive, difficult to
standardize, time consuming
a) Ascheim-Zondek PT
(+): corpus luteum formation induced by B-hCG in
prepubertal MICE
b) Friedman PT
(+): ovulation in mature female RABBITS
c) Frog Test
(+): ovulation in mature female FROG
2. AGGLUTINATION IMMUNOASSAY
-

Limitations: less sensitive and specific; longer waiting

time
a) Hemeagglutination Inhibition Test (HAI)
b) Latex agglutination inhibition test (LAI)
c) Direct Latex particle agglutination test
# 1-2: Indirect
(+) ABSENCE of AGGLUTINATION
#3: Direct
(+) PRESENCE of AGGLUTINATION
sensitivity: >500 mIU/ml

Difference between hemeagglutination & latex

agglutination inhibition test

3. RADIOIMMUNOASSAY
- Sample: serum
- 2 types based on anti-sera used:
B-sub-unit RIA
-Specific for B-hCG
-Sensitivity: 5 mIU/ml
-Unaffected by increased levels of LH
Whole hCG RIA
-Detects both alpha and beta subunits hCG
-Affected by high levels of LH; less specific (BFP)
4. ENZYME-LINKED IMMUNOSORBENT ASSAY (ELISA)
- A double monoclonal Ab technique
- As specific for B-hCG subunit RIA
- Sample: urine
- Sensitivity: >50 mIU/ml
- Ex: Abbotts TEST Pack
5. RADIORECEPTOR ASSAY
- Used in reference laboratories only
- Utilizes receptors from ovaries of pregnant cows
- Sensitivity: >5 mIU/ml
- Prone to BFP by LH
- Sample: serum or urine
CLINICAL APPLICATIONS OF PT
A. DIAGNOSIS OF PREGNANCY
- Levels of hCG in pregnancy = days/wks AOG
- Serum level of:

hCG levels increase 2x q 2 days during the first 6 wks

AOG
o Serum hCG reaches its detectable level of 5mIU/ml
within the 1st 24 hrs from the time of implantation
o 25 mIU/ml --- 8-10 days after conception (22-26 d from
LMP)
o 500 mIU/ml --- 14-18 days after conception (28-32 d
from LMP)
o peaks to 20,000 57,000 mIU/ml --- 55-70 d after
conception
o rapidly declines to a steady level of 10, 000 mIU/ml
after the first trimester of the pregnancy.
o Undetectable: within 14 days postpartum
o 1st wk AOG: serum hCG is> urine hCG
o >3rd wk AOG: urine hCG > serum hCG
- Sensitivity:
o RIA & ELISA: detects pregnancy at the time of the first
missed period
o HAI: 14 days after the missed period
o LAI: 21 days after the first missed period
B. ECTOPIC PREGNANCY
- Extra-uterine pregnancy
- hCG level: 500-800 mIU/ml
- Lower than normal pregnancy
- An adjunct test only
- Correlate with: CM, PE, UTZ findings
C. THREATENED ABORTION
- Impending abortion
- hCG levels will either show:
NO increase
Progressive decline
D. TROPHOBLASTIC TUMORS
- Both usually present with markedly elevated hCG
sometimes reaching 6M/ml or persistently increaseing
hCG levels
o Hydatidiform mole
o Germ Cell tumors (ex: Choriocarcinoma)
- For H mole: serum B-hCG level is monitored even after
evacuation of H. mole to detect recurrence
o

After evacuation of H. mole: hCG levels should be

undetectable within 2-3 mos because of the possibility of
H. mole to develop into choriocarcinoma
OTHER PREGNANCY RELATED EXAGGERATED INCREASE
IN hCG:
- Multiple pregnancy
- Polyhydramnios
- Exclampsia
- EBF
E. TESTICULAR TUMORS
- Increase level of hCG in the following tumors:
Seminomas
Teratomas
Embryonic carcinoma
- Mandatory to perform hCG levels on these cases, if
increased hCG levels can be used as a tumor marker for
prognostication and monitoring of treatment
NON-TROPHOBLASTIC TUMORS
- Not routinely done because it is not specific
- Seen in:
o Gastric Carcinoma
o Hepatoma
o Pancreatic carcinoma
o Lymphoma
o leukemia
-

URINARY CALCULI ANALYSIS

Formation and Physiology
Factors that predispose to calculi formation:
1. metabolic, nutritional, or idiopathic disturbances (ex:
gout)
2. Endocrinopathy (ex: hyperparathyroidism)
3. Urinary obstruction
4. Infection with urea-splitting organism
5. Mucosal changes (ex: dietary deficiency)
6. Extrinsic and environmental factors (DHN, alkali
excess, chemotherapy)
7. pH
<5.5: uric acid, cystine or xanthine calculi
5.5-6.0: calcium oxalate and apatite calculi

>7.0: Mg ammonium phosphate and calcium

phosphate calculi
Classification
a. based on location
b. chemical nature
Calcium calculi= most common
Urinary calculi analysis
Physical Examination:
1. Size
2. Shape
3. Color
4. Texture
5. Internal structure (laminations)
Composition analysis:
1. Chemical analysis
2. Polarizing microscopy
3. Radiographic diffraction
4. Infrared spectroscopy
5. Electron microscopy
Causes of various calculi
1. Calculi of calcium composition:
a. Idiopathic hypercalciuria
b. Primary hyperparathyroidism
c. bone diseases
d. excessive milk, alkali, vit D
e. Renal tubular disease
f. sarcoidosis
g. Berylliosis
2. Calculi of calcium oxalate
a. oxaluria
b. incomplete catbolism of carbohydrates
c. isohydria at ph 5.5 6.0
d. excessive glycogen breakdown
3. Calculi of calcium phosphate composition
a. same causes as for Ca oxalate
b. alkaline infection
c. persistently alkaline urine
4. Calculi of magnesium ammonium phosphate
composition
a. alkaline infection
5. Calculi of uric acid and urate

a. gout
b. polycythemia
c. leukemia
d. lymphoma
e. liver disease
f. acid isohydria
g. theophylline and thiazide tx
h. rapid protein catabolism
6. Calculi of cystine composition
a. transient acute phases of chronic renal dses
b. heavy metal nephrotoxicity
c. aminoaciduria
d. renal tubular acidosis syndromes

CEREBROSPINAL FLUID ANALYSIS

Formation & Physiology:
- Produced by the choroid plexi by filtration under
hydrostatic pressure, reabsorbed by the arachnoid villi &
flows at the subarachnoid spaces
- Rate of production:
20 ml/hr or 500 ml/day
- Total volume: neonates:
10 60 ml
Adults:
140 170 ml
- Distribution: Ventricles: 20 ml
Subarachnoid spaces: 60 ml
Spinal canal: 80 ml
- Functions:
o supply nutrients
o remove metabolic waste products
o mechanical barrier
Specimen Collection & Handling
Methods of collection:
a. Lumbar tap: 3rd, 4th,5th lumbar vertebrae
b. Subdural tap
Sampling:
- Three sterile bottles
1st = chemical and serological tests (freeze)
2nd = microbiological study (room temp)
3rd = cell count (refrigerate)
- Done as STAT
- 2-4 ml/bottle
Routine CSF Analysis

PHYSICAL EXAMINATION
1. CSF pressure
- Normal : 100-200 mm H2O
- Increased in:
o meningitis
o subarachnoid bleeding
o brain tumor/abscess
- Queckenstedt Sign = a sign produce when jugular vein
compression is done causing increase in venous pressure
and eventually an increase in CSF pressure at the lumbar
area. If there is a subarachnoid block above the lumbar
area, the Queckenstedt sign is negative.
2. Appearance and color
- Normal: clear and colorless
- NB: A CLEAR CSF DOES NOT rule out INTRACEREBRAL
BLEED because RBC = <360/cubic mm will not give a
visible bloody sample!
3. Clot formation
- Clot formation + blood = traumatic tap
- Clot formation but no blood = meningitis, Froins
syndrome, blockage of CSF Pathway
- Web-like clot, no blood = TB meningitis
CHEMICAL EXAMINATION
1. Glucose determination
- NV: adults:60-70% of plasma glucose (45 mg/dl or 2.5
mmol/L)
neonates: 80% of plasma glucose
- must be tested together with blood glucose which is done
2 hrs prior to the lumbar tap because it takes 30 mins to 2
hrs before changes in blood glucose is reflected in the CSF
- Glucose Clinical significance:
o Decreased:
bacterial, fungal, tubercular,
amebic, leptospital meningitis, metastatic Ca,
subarachnoid bleed hypoglycemia
o Increased: DM, IVF tx
o Normal: Early stage of any type of meningitis, viral
meningitis, encephalitis, brain tumor
o Methodology: colorimetric; enzymatic
2. Protein Analysis of CSF
- NV:
6 mos to adults:
15-45 mg/dl

Newborn: 75 150 mg/dl

>60 y/o: > 60 mg/dl
- Fractions:albumin (major)
pre-albumin
alpha globulins (haptoglobulin, ceruloplasmin)
beta globulin (transferin)
tau (CHO-free transferin; seen ONLY in CSF)
gamma globulin (IgG and IgA)
- NB:
CSF has NO fibrinogen and IgM!
- Clinical significance of protein detn:
o Increased:
damage to blood-brain barrier
production of Ig in the CNS
degeneration of neural tissues
traumatic tap (1 mg CHON=1000 rbc)
o Decreased: CNS leakage
- Albuminocytologic dissociation
= increase CHON, normal cell count
= seen in Guillain-Barre syndrome
- Oligoclonal bands ( > 2)
= Increase Ig = Multiple sclerosis (MS),
GBS
- Myelin Basic Protein (MBP)
= demyelination= MS
Important Indices
- CSF-serum albumin ratio
o <9 = intact BBB
o >9 = damaged BBB
- CSF-serum globulin ratio
o 0.77 = production of Ig within the CNS
Methodology:
a. Pandys Test - a bedside semi-quantitative test for
protein done by adding PHENOL to the CSF sample;
precipitates out mostly globulins; obsolete
b. Colloidal Gold Test - for globulin
- once used in detecting MS and Tabes dorsalis
- Dye-binding mtd; obsolete; not specific & sensitive
c. Turbidimetry (SSA or TCA)
d. Immunoassay more specific and sensitive
- Electrophoresis, RIA, Nephelometry
3. CSF Lactate
- Normal: 25 mg/dL

Clinical significance: >35 mg/dL = bacterial meningitis

severe head injuries
>25 mg/dL = tubercular and fungal
meningitis
< 25 mg/dL = viral meningitis
4. CSF glutamine
- Normal: 8-18 mg/dL
- Clinical significance:
o increased: hepatic encephalopathy
Reyes syndrome
coma
5. CK-BB and LDH
o increased: post cardiac arrest
poor prognosis
-

CELL COUNT
1. WBC count
- Normal: adult: 0-5 /uL NB: up to 30/uL
- corrections for contamination:rbc: 700 rbc = 1 wbc
2. Differential count
- Normal:predominantly lymphocyte
- Clinical significance: see table
MICROBIOLOGIC STUDY
1. Gram stain
- Most common microbes:
a. Strep. Pneumoniae
b. Hemophilus influenzae
c. Escherichia coli
d. Neisseria meningitidis
e. Listeria monocytogenes
Limulus lysate test = (+) Gram (-) bacteria
Most Common Cause of Bacterial Meningitis according
to Age Group:
1 3 mos
B-Streptococcus (Strep. agalactiae)
Escherichia coli
Listeria monocytogenes
Enteric Gram (-) bacilli
3 mos 6 y/o
Haemophilus influenzae
Neisseria meningitides
(Meningococci)
Streptococcus pneumoniae

Older children
Meningococci
to adolescents
Streptococcus pneumoniae
(Pneumococci)
Adults
Meningococci
Streptococcus pneumoniae
Old Age
Pneumococci
Meningococci
Gram (-) bacilli
2. Acid Fast Bacilli stain = for TB meningitis
3. India Ink prepn = for Cryptococcus neoformans
4. Culture and Sensitivity testing = identify causative
agents in meningitis
SEROLOGIC TESTS
1. ELISA methods for detecting:
a. Streptococcus Group B
b. Haemophilus influenzae type B
c. Streptococcus pneumoniae
d. Neiserria meningitides A,B,C,Y,W135
e. Escherichia coli K1
2. Latex Agglutination Test for Cryptococcus
neoformans
3. Limulus Lysate Test for Gram (-) bacilli
- Principle: The endotoxin produce by the walls of Gram (-)
organisms will coagulate the amebocyte lysate within 1 hr
if incubated at 37 C.
4. Neurosyphillis Tests
a. VDRL (Venereal Disease Research Laboratory Test) least specific
b. RPR (Rapid Plasma Reagin Test) - simplest; least
sensitive & specific
c. FTA-ABS (Fluorescet-Treponemal Antibody-Absorption
Test) - most specific and sensitive

SEMINAL FLUID ANALYSIS

Formation & Physiology
1. Seminal fluid
4 fractions:
a. testes & epididymis (5%) = storage of sperm cells
b. seminal vessicles (60%) = fructose = nutrient; motility

c. prostate gland (20-30%) = ACP, citric acid, zinc, proteolytic

enzymes
= coagulation and liquefaction
d. bulbourethral glands (5%) = thick alkaline mucus
= neutralizes vaginal and prostate secretions
2. Spermatozoa = seminiferous tubules of the testes
Specimen Collection & Handling
Collection:
- mtd: masturbation
- coital fasting for 3-5 days
- 2 wks interval for evaluation of infertility
- warm container
- examined within 1 hr from collection
- potential reservoir for HIV and Hepa virus
ROUTINE SEMEN ANALYSIS
A. Physical Examination
1. Coagulation and liquefaction
- normal: clotted upon ejaculation; liquefies 30-60 mins
after
- abnormal: failure to clot/liquefy
= low prostatic enzymes = infertility
2. Volume
- normal: 2-5 ml
- abnormal: decreased = infertility
3. Appearance
- Normal: gray-white, translucent
- Abnormal: white = infection
red = trauma
yellow = prolonged abstinence; urine
contamination, medications
4. Viscosity
- Normal: pour in droplets
- Abnormal: clumped and highly viscous= incomplete
liquefaction = infertility
5. pH
- Normal: 7.2 8.0
- Abnormal: >8 = infection
acidic = decreased prostatic secretions
B. Sperm Count
- Normal:740 M/ejaculate
20-160 M/ ml
- Clinical significance:

- valid measurement of infertility

- Borderline: 10-20 M/ml
- < 10 M/ml = infertility
C. Sperm Motility
- Normal motility: forward progressive movement
50% grade 2 after 1 hr
Grading:
4
rapid, straight-line motility
3
slower movt, some lateral movt
2
slow forward movt, noticeable lateral movt
1
no forward progression
0
no movement
- Methodology:
1. manual
2. Computer-assisted semen analysis (CASA)
= measures both sperm velocity and trajectory
3. Eosin-nigrosin staining
viable = does not take the stain
dead = take up the stain
4. Antisperm Antibody (in men and women)
a. Mixed Agglutinatio Reaction test = detect
presence of
male sperm antibodies IgG)
Normal: <10% agglutinated sperm
b. Immunobead test= detect prsence of female
antisperm
antibodies (IgM, IgG, IgA)
Normal: < 20% agglutinated sperm
D. Sperm Morphology Study
- Normal morphology:
= oval head, short neck and long tail
= > 30% (strict criteria); >50% (routine criteria)
- Abnormal morphology:
a. head abnormality
= poor penetration of ovum
= double, pin, amorphous, constricted, giant
b. tail abnormality = poor motility
c. Neck abnormality = head bend
- Krugers criteria = strict measurement of head, neck
and tail; recommended by WHO
-

E. Chemical tests

Normal Semen Chemical Values

1. fructose
> 13 umol/ejaculate
2. neutral a- glucosidase > 20 mu/ejaculate
3. Zinc
> 2.4 umol/ejaculate
4. Citric acid
> 52 umol/ejaculate
5. ACP
> 200/ejaculate
- Clinical significance:
decreased fructose = lack of seminal vesicle fluid
decreased 25 = lack of prostatic secretions
F. Microbiologic Tests
- WBC count = 1 M/ml = infection (prostate)
= requires C & S
- Common microbes isolated:
Chlamydia trachomatis
Mycoplasma hominis
Ureaplasma urealyticum
Enterobacter
Anaerobes
G. Medicolegal seminal test
- ID of sperm cells
- Test for ACP
- ABO typing
- DNA testing
H. Postvasectomy Semen Analysis
- check completeness of sterilization
- Test: presence or absence of sperm cells
- done at monthly interval 2 mos after vasectomy
- endpoint: two consecutive monthly specimens show
azospermia
I. Sperm Function test
- done in advance Andrology laboratories
- Hamster Egg penetration
- Cervical mucus penetration test
- Hypo-osmotic swelling
- In vitro acrosome reaction
-

Test
Appearance
green/bilous
peritoneal lavage
w/

Clinical significance
GB, pancreatic disorders
Blunt trauma injury

>100,000 rbc/uL
WBC >500/uL
CEA
CA 125
Amylase

Bacterial peritonitis, cirrhosis

Malignancy (GIT)
Malignancy (ovarian)
Increased in pancreatitis, GIT
perforation
Increased in perforation in GIT

ALP
BUN/crea
Gram stain; C & S

Ruptured or punctured urinary

bladder
Bacterial peritonitis

AFB stain

TB

Adenosine
deaminase

TB

SYNOVIAL FLUID ANALYSIS

Formation & Physiology:
- fluid found in movable joints (diarthoses)
- provides nutrients, serve as cushion ultrafiltrate of plasma
form by nonselective filtration across the synovial
membrane
Specimen collection and handling:
- Mtd of collection: Arthrocentesis
3 tubes:
1 = heparinized: microbio
2 = EDTA: hema tests
3 = plain: other tests
Routine Synovial Fluid Analysis
- Macroscopic:
a. Volume: <3.5 ml
b. Color and clarity: pale yellow & clear
c. Viscosity: (+) string test 4-6 cm long
(hyaluronidase)
d. Clot formation: none
- Microscopic:
a. Rbc:
<2000 /uL
b. WBC: < 200 /uL
c. Neutrophils: < 20% of differential count
d. Lymphocytes: <15 % of diff count
e. Monocytes & macrophages: 65% of diff ct

f. Crystals; none
Chemical tests:
a. Glu: <10 mg/dL lower than blood glucose
b. Lactate: <250 mg/dL
c. Total protein: <3g/dL
d. Uric acid: = blood level
- Microbiologic:
a. Gram stain, AFB stain, KOH
b. C and S
- Serologic tests:
Ab for SLE, RA, Lyme dse (Borrelia burgdorferi)
Clinical significance (Abnormal findings)
Macroscopic findings:
- (+) Clot formation = fibrinogen, hemorrhage, damaged
synovial membrane
Microscopic findings:
a. increased rbc - hemorrhage, traumatic tap
b. WBC >2,000/uL with increased PMN - bacterial
arthritis, acute gouty arthritis rheumatoid arthritis
c. increased eosinophils - rheumatic fever, parasitic
infection
d. crystals
- monosodium urate monohydrate = gouty arthritis
- calcium pyrophosphate dehydrate = pseudogout
- cholesterol = rheumatoid arthritis
Chemical findings:
a. Decreased glu = inflammatory/sepsis
b. Increased protein = damaged synovial membrane
c. Increased uric acid = gout
d. Increased lactate = septic arthritis
Except: Gonococcal arthritis
Microbiologic:
common isolates:
a. Hemophilus
b. Neisseria gonorrhea
c. Staphylococcus
d. Streptococcus
e. fungal, TB
-

GASTRIC FLUID

Physiology of Gastric secretion and digestion

a. stomach = reservoir for ingested food
b. Initiation of protein digestion by pepsin
c. Secretes intrinsic factor for binding and absorption of
B12 in the ileum
3 General Phases of Gastric secretion:
A. Cephalic phase
- stimulates gastric secretion
a. Gastrin secretion by G-cells and d-cells
b. HCl secretion by parietal cells
c. Pepsin secretion by chief cells
- Gastric peristalsis and emptying is promoted
B. Gastric phase Continued Gastric distention and
secretion
C. Intestinal phase
- Digestion in the small intestine
- Simplified food (glu, amino acids, fats) stimulate secretion
of Gastric inhibitory substance (GIP) from the duodenum
- inhibits gastrin secretion
Chemical composition
1. HCl acid
- parietal cells (fundus)
- converts pepsinogen to pepsin
- hydrolize polypeptides and disaccharides in the presence
of dissacharidases
2. Pepsin - catalyzes proteolysis
3. Mucus - protect gastric mucosa from autodigestion
4. Miscellaneous substances - enzymes, proteins,
intrinsic factor
Gastric analysis and collection
A. Basal gastric secretion test
- measures 15 hr fasting levels of gastric production
- four 15-30 min interval specimen is collected and the ff
tests are done:
a. Volume
b. pH
c. titrable acidity
d. calculated acid output
B. Maximal stimulation tests
- uses histamine or pentagastrin to stimulate
maximal
secretion
C. Insulin-induced hypoglycemia

measures completeness of vagotomy

insulin induced hypoglycemia (<50 mg/dL) stimulates
gastrin secretion after 2 hrs
D. Tubeless gastric analysis
- non-invasive
- uses a dye (dianex blue)
Routine Examination
appearance: translucent
Volume: 50-75 ml
Odor: faintly pungent
Mucus: quantity varies
Measurement of gastric acid and pH
Microscopic
Clinical significance
- Generally for the diagnostic of digestive disorders or
ulcers
-

Categories of gastric acidity:

o anacidity: failure of gastric acidity to fall lower than 6.0
in a stimulation test
o hypochlorhydria: physiologic failure of ph to fall lower
than 3.5, but decreases 1.0 pH or more upon gastric
stimulation
o achlorhydria: physiologic failure of ph to fall lower than
3.5 even with gastric stimulation
Pre-operative procedure to know extent of surgery for
PUD
diagnosis of Zollinger-Ellison syndrome = pancreas
malignancy causing increase gastric acid and gastrin
secretion
Assessment of completeness of vagotomy