Amniotic fluid,h cg, SPUTUM & BRONCHOALVEOLAR LAVAGE, sweat, test for HDN, Hemolytic Disease of the Newborn, Fetal Lung Maturity,

3. PRIMARY FUNCTIONS
1. Provide cushion for
the fetus
2. Allow fetal movement
AMNIOTIC FLUID VOLUME
• From fetal urine and lung
fluids
• After the first trimester,
FETAL URINE is the major
contributor of the
amniotic fluid volume
• approximately 35 mL
during the 1st trimester,
peaks during the 3rd
trimester (approx. 1 L)

4. POLYHYDRAMNIOS
 INCREASED amniotic fluid
volume
CAUSES:
 Decreased fetal swallowing of
urine
 Fetal distress; Neural tube
defects
OLIGOHYDRAMNIOS
 DECREASED amniotic fluid
volume
CAUSES:
 Increased Fetal swallowing
of urine
 Membrane leakage
 Urinary Tract Deformaties
SPECIMEN COLLECTION!
METHOD OF COLLECTION  AMNIOCENTESIS –transabdominal
Up to 30mL (max) is collected in sterile syringe
 Performed after the 14th week of gestation
2ND TRIMESTER AMNIOCENTESIS (4-6mos)  Assess genetic defects
3RD TRIMESTER AMNIOCENESIS (7-9mos)  Fetal Lung Maturity (FLM)
 The fluid is dispensed into sterile plastic specimen containers

5. SPECIMEN HANDLING!
Test for Fetal Lung Maturity (FLM) Place on Ice (delivery)
Refrigerated or Frozen up to 72 hours prior to
testing;
Filtration or low speed centrifugation is
recommended to prevent loss of phospholipids
Test for Cytogenetic Studies Room temperature or body temp
Test for Hemolytic Disease of the
Newborn (Bilirubin)
Protect from light
placed in amber bottles
Test for Chemistry Separated from cellular elements and debris ASAP
If need to be stored more than 24 hrs-frozen

6. ANALYTE AMNIOTIC FLUD MATERNAL URINE
LESS RELIABLE
PROTEIN + O
GLUCOSE + O
MORE RELIABLE
UREA <30 mg/dL >300 mg/dL
CREATININE <3.5 mg/dL >10mg/dL
AMNIOTIC FLUID VS MATERNAL URINE
FERN TEST!
Specimen (Vaginal Fluid)
Slide (Air Dry)
(+) Fern-Like crystals- Amniotic
Fluid (Screening)

7. A.K.A O.D. 450
Principle: Spectrophotometry
Absorbance of amniotic fluid
NORMAL  Increase at 365nm, decrease at 550nmm
(<0.025 )
HDN  Increase at 450 nm
Results are plotted on a LILEY GRAPH:
ZONE I = Nonaffected/ mildly affected fetus
ZONE II = Moderately affected fetus (requires close
monitoring)
ZONE III = Severely Affected fetus (requires
intervention)

12. • Reported as MoM (Multiples of the Median)
 Spinda Bifida  open defect
 Anencephaly  brain stem only
SCREENING TEST  Alpha-Feto Protein (AFP)
Principle: Immunoassay
NV: <2.0 MoM
o Increased in NTD
o Decreased in Down Syndrome
CONFIRMATORY TEST  Acetylcholinesterase
Principle: Spectrophotometry
NV: undetectable

13. TEST PRINCIPLE INFORMATION NORMAL
VALUE
L/S ratio Thin-layer
chromatography
Reference Method
Lecithin  for alveolar stability
(phopholipid)
Sphingomyelin  serves as control
*cannot be done with specimen
with contaminated blood or
meconium
>2.0
Amniostat-FLM Agglutination
immunoassay
Immunologic test for phosphatidyl
glycerol(Production is delayed
among diabetic mothers)
Not affected by blood or by
meconium
POSITIVE
Foam Stability Index Modified foam-
shake
95% ethanol used as anti-foaming
agent
(+) Foam/ Bubbles = Mature Fetal
Lung
≥47

14. TEST PRINCIPLE INFORMATION NORMAL
VALUE
Microviscosity Fluorescence-
polarization
Albumin used as internal standard
The presence of phospholipids
decreases microviscosity
≥55 mg/g
Lamellar body count Resistance pulse
counting
Lamellar Bodies (aka Type II
pneumocytes)
 Responsible for alveolar
surfactants production
Uses the platelet channel of
hematology analyzers
≥32,000/
mL =
adequate
maturity
OD at 650 nm Spectro
photometry
Requires centrifugation at 2000 g for
10 min
*Increased Lamellar bodies =
Increased OD (Absorbance)
An OD of >0.150 is equivalent to:
 L/S ratio of >2.0
 (+) Phosphatidyl glycerol
≥0.150

15. RESPIRATORY DISTRESS SYNDROME (RDS)
 Most frequent complication of early delivery TEST FOR FETAL AGE
CREATININE!
* >2.0mg/dL amniotic
fluid creatinine = 36
weeks (9mos)
TEST FOR DETECTING PRE-
TERM DELIVERY:
FETAL FIBRONECTIN
 between week 22 and
week 34 of pregnancy

17. Produced by the CYTOTROPHOBLASTS CELLS
of the Placenta
Peaks during 1st Trimester of pregnancy
(Increased in Blood, Urine & Amniotic Fluid)
Composed of two subunits:
Alpha = hCG, LH, FSH, TSH
Beta= confers specificity for the hCG
HOME-BASED hCG PREGNANCY KIT
Principle: Enzyme Immunoassay
Specimen: First Morning Urine

18. 1. Immunoassays
 Principle: Detection of hCG using monoclonal antibodies.
 Methods:
1) Agglutination Immunoassays – direct or
agglutination-inhibition
2) Competitive Radioimmunoassay
a) Principle: serum hCG and the radiolabeled hCG
compete for the binding of anti-hCG
b) Sensitivity – 5 mIU/Ml
3) EIA (Sandwich ELISA)
a) Principle: Detection of hCG based on color
indicator reaction mediated by an enzyme (e.g.
ALP); commonly used in home-based pregnancy
tests
b) Sensitivity – 2-5 mIU/mL

19. 3) Immunochromatography (lateral flow tests)
a) Principle: The labeled antibody-dye conjugate in the
reaction zone binds to the hCG in the specimen forming an
antibody-antigen complex. This complex binds to the anti-
hCG antibody in the test zone and produces a colored band
when the hCG concentration is equal to or greater than 20
mIU/ml. In the absence of hCG, the reaction mixture
continues flowing through the absorbent device past the
test zone allowing the binding of unbound conjugates to the
reagent in the control zone.
b) Interpretation of Results:

20. 2. Bioassays
SOURCES OF ERROR!
1. False-positive: production of hCG in the pituitary; tumors characterized by
significant amounts of hCG; ectopic pregnancy and incomplete abortion;
intake of chlorpromazine, phenothiazine, and aspirin
2. False-negative: low titer or concentration of hCG; low sensitivity of test
animal or assay method; use of toxic urine (bacterial contamination, increased
electrolyte levels, salicylates, and barbiturates)

22.  From Upper & lower respiratory tract (not sterile)
 Tracheobrochial secretions (mixture of plasma, electroytes,
mucin & water) added with cellular exfoliations, nasal &
salivary gland secretions and normal oral flora
 STORAGE: refrigerate or formalin
SPUTUM COLLECTION!
Expectoration-FIRST MORNING MOST PREFFERED (ROUTINE)
24-HOUR For volume measurement
THROAT SWAB For Pediatric patients
TRACHEAL ASPIRATION For delibitated patients
SPUTUM INDUCTION For non-cooperative patients
BRONCHOALVEOLAR LAVAGE infusion of saline through a
bronchoscope followed by aspiration

23. TRACHEAL
ASPIRATION
SPUTUM INDUCTION
MACROSCOPIC EXAMINATION
VOLUME DECREASED: Bronchial asthma, acute bronchitis, early pneumonia,
stage of healing
INCREASED: Bronchiectasis, Lung Abscess, edema, gangrene,
tuberculosis, pulmonary hemorrhage
ODOR ODORLESS  Normal
Foul or Putrid  Lung gangrene, advanced necrotizing tumors
Sweetish  Bronchiectasis, Tuberculosis
Cheesy  Necrosis, Tumors, empyema
Fecal liver abscess, enteric Gram negative bacterial infection

24. MACROSCOPIC EXAMINATION
COLOR Colorless Or Translucent  Made of mucus only
White or Yellow /Yellow Green Increase Pus (TB, Bronchitis,
jaundice, pneumonia)
Gray increase pus & epithelial cells
Bright green  jaundice, caseous pneumonia, Pseudomonas infection,
rupture of liver abscess
Red/bright red  recent hemorrhage (acute cardiac or pulmonary
infarction, neoplasm invasion) ,TB, Brochiectasis
Anchovy sauce or Rusty Brown  decomposed hemoglobin (lobar or
pneumococcal pneumonia, pulmonary gangrene)
Prune Juice  Pneumonia, Chronic Lung cancer
Olive green/grass green – chronic cancer
Black  Inhalation of dust particles, carbon or charcoal, heavy
smokers, anthracosis
Rusty with pus  Lobar Pneumonia
Rusty without pus  Congestive Heart Failure
Currant, jelly-like  Klebsiella pneumonia infection
CONSISTENCY MUCOID  asthma, bronchitis
SEROUS OR FROTHY  lung edema
MUCOPURULENT  Brochiectasis, TB with cavities

25. MACROSCOPIC STRUCTURES
CLINICAL
SIGNIFICANCE
Dittrich’s plugs yellowish or gray caseous materials
about the size of a pinhead that give a foul odor
when crushed
Bronchiectasis
putrid bronchitis
bronchial asthma
Pneumoliths/
Broncholiths /Lung
stones
small white or gray fragments from the
calcification of infected and necrotic tissue within
the bronchial cavity
Hard concretion in a bronchus
chronic PTB
histoplasmosis
Bronchial casts white or gray branching tree-like casts from the
bronchioles
lobar pneumonia
fibrinous bronchitis
diphtheria
Cheesy masses fragments of necrotic pulmonary tissue that range
in size from pinpoint to pea-size
pulmonary gangrene
pulmonary TB
lung abscess
Mycetomas rounded masses of fungal elements  Aspergillus infection
Layer Formation 3 layers:
1. 1st (top) = frothy mucus
2. 2nd (middle) = opaque, water material
3. 3rd (bottom) = pus, bacteria, tissues
Bronchiectasis
lung abscess
 gangrene
Foreign Bodies  Bronchial calculi (made of calcium carbonate &
phosphate)
 Asbestos bodies, silica partciles (dust partcles
in BAL)
Pneumoconiosis

26. MICROSCOPIC STRUCTURES
CLINICAL
SIGNIFICANCE
Elastic fibers Slender fibrils with double contour and curled ends
refractile fibers shed off during the cougning out
process; indicates destructive disease
Tuberculosis
Charcot-Leyden
Crystals
Colorless hexagonal, needle-like or bipyramidal
crystals
Arise from disintegration of eosinophils
 Bronchial asthma
Pigmented cells Heart failure cells  Hemosiderin-laden macrophages
Carbon-laden cells  angular black granules
Congestive Heart Failure
Heavy Smokers
pneumoconioses
Curschmann’s
spirals
 spirally twisted mucoid strands frequently coiled into
little balls
Bronchial asthma
Myelin Globules Colorless globules occuring in a variety of sizes and
bizzare forms
No CS but maybe
mistaken as Blastomyces
Epithelial cells Creola bodies  Clusters of bronchial epithelial cells
with vacuolated cytoplasm and ciliated borders
Bronchial asthma
Fungi  C. Albicans, C. Neoformans, C. Immitis,
H.capsulatum, B. Dermatitidis, A. fumigatus
Parasites  Migrative larva: (ASH)
 E. Histolytica, E.gingivalis, T.tenax, P. Westermani
(operculated egg), E. Granulosus, T. canis

27. Charcot-Leyden Crystals
(Heart failure cells )
Hemosiderin-laden macrophages Curschmann’s spirals
Expectorated Bronchial cast
Layer Formation
Broncholiths

28. • Important diagnostic test for Pneumocystis carinii (P.
Jiroveci) in immunocopromised patients
• MACROPHAGE  MOST PREDOMNANT CELL SEEN

29. SWEAT TEST
 Used to diagnose Cystic Fibrosis
(Mucoviscidosis);
Cystic Fibrosis
autosomal recessive disorder
Metabolic disease that affects the mucous
secreting glands of the body
Associated with pancreatic insufficiency,
respiratory distress & intestinal obstruction
Gibson and Cooke PILOCARPINE
IONTOPHORESIS
Pilocarpine + mild current =stimulates
sweats glands
Sweat is tested for Sodium and Chloride
SWEAT Na &
Cl Values:
Diagnostic for
CF = >70meq/L
Borderline for
CF = 40meq/L

31. List of References
Lillian Mundt & Kristy Shanahan, Graff’s
Textbook of Urinalysis and Body Fluids, 2nd Ed.
Susan Strassinger & Marjorie Di Lorenzo,
Urinalysis and Body Fluids, 5th & 6th Ed.
Erol Coderres,RMT-AUBF notes
Roderick Balce, RMT-CEU Professor AUBF
Notes