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Clinical Pathology

Practical 2
Review

Special thanks to Pat Siao

Acetone Powder:
Sodium nitroprusside
Ammonium sulfate
Sodium carbonate,
anhydrous

Detection of ACETONE in urine using Acetone Powd

Detection of Acetone in urine using


Acetone Powder

No violet color =
(-) ACETONE in urine

VIOLET color =
Presence of ACETONE in
urine
(+) Ketosis or Diabetes

Analysis #21: Quantitative Determination of


Total PROTEIN

Tests for: Incipient Renal Disease


Pyrogallol red-molybdate complex (absorbance 467nm)
PRM + protein purple color inc. absorbance (598nm)

Quantitative Determination
of Protein

Au
Tp
Cs vol (dL)
As

Example Practical Question:


Au = .20
As = .10
Cs = 50 mg/dL
Urine Volume = 10 dL
= 1000 mg/24 hrs or 1g/24 hrs
Normal = 0.28 1.41 g/24 hrs

Analysis #22: Micral Test


Immunological detection
of human albumin (IgG)
Absorbed urine on strip
binds antibody-goldconjugate to urine
albumin
Color produced is directly
related to albumin content
Ideal specimen: First
morning urine

(+) Positive when 2/3 morning urine specimens > 20


mg/L = persistent microalbuminuria
(-) negative < 20 mg/L = physiologic urine albumin

Analysis #23: Blood Urea Nitrogen (Modified


Berthelot)

Urea Urease
CO 2 Ammonium
Ammonium hypochlori
te & nitroferri
cyanide
Chloramine
Chloramine Salicylate
Quinonechloramine
Quinonechloramine Salicylate
indophenol chromophore (630nm)

Analysis #23: Blood Urea Nitrogen (Modified


Berthelot)

Au
BUN
Cstd
As
Example Practical
Question:
Au = .20
As = .10
Conc. Of Standard = 25
mg/dL
= 50 mg/dL (high

High BUN

Low BUN

Impaired
kidney
function

Liver disease

Kidney
disease

Malnutrition

dehydration

overhydratio
n

Analysis #25: Creatinine


Clearance
Unknown:
1 mL urine
1.5 mL 10% NaOH
20 mL picric acid
77.5 mL distilled
water

Standard:
1 mL Creatinine
Std
1.5 mL 10% NaOH
20 mL picric acid
77.5 mL distilled
water

Tests for: kidney function


Specimen: 24 hour urine specimen
Creatinine + Picric acid forms red color proportional to amount of
creatinine

Analysis #25: Creatinine


Clearance
RU
Urine Creatinine
Cstd
RS
Example Practical Question:
RU = .15
RS = .10
Conc. Of Standard = 100 mg/dL
= 150 mg/dL

Analysis #25: Creatinine


Clearance
Urine Creatinine (mg/dL) x vol. of urine (cc/min)
Creatinine Clearance
Serum Creatinine (mg/dL)

Example Practical
Question:
Urine Creatinine = 200
mg/dL
Serum Creatinine = 2
mg/dL
Total Volume = 2 cc/min

Microscope Findings in
Urine

Uric Acid Crystals

Renal stones, gout, high purine metabolism, acute febrile conditions,


chronic nephritis

Calcium Oxalate Crystals

Renal stones, DM, liver disease, chronic renal disease, can be found
in normal individuals

Triple Phosphate

Renal Canaliculi, Chronic Pyelitis, Enlarged prostate, UTI

Hyaline Cast

Severe renal disease with increased numbers, healthy individuals


after heavy exercise

Granular Cast

Nonpathologic strenuous exercise/stress;


Glomerulonephritis, Pyelonephritis

Pathologic

Bacteria

With increased WBC UTI

Squamous Epithelial Cells

Occur in urethra and vagina; Vaginal contamination

Yeast Cells (Budding Yeast)

UTI especially from diabetic parents, immunosuppressed patients,


skin or vaginal inefections

Amorphous Urates

Pus Cells

RBC

Can originate from any part of the urinary tract, Glomerulonephritis,


Trauma, Renal diseases

Analysis #27
Hema-screen Slide Test

Tests for: fecal occult blood


Hemoglobin comes in contact with guaiac.
Peroxide developing solution guaiac-peroxidase
reaction occurs making blue-green color

(+)
Normal in
Infants
Abnormal in
Adults
Positive
(+)
Any trace of
blue color,
within or on
the outer rim
of the
specimen, is
positive for
occult blood.

(-)
Abnormal in
Infants
Normal in
Adults
Negative
(-)

Infants:
GI is sterile
transit time
due to shorter
bowel
(-) =
Abnormal
Obstructi
on

nalysis #28: Neutral Fat Estimation (Sudan


III Stain)

Neutral Fat Droplet


Indicated by yellow-orange red
globules

NORMAL
< 50 fat
globules/hpf

Negative
(-)
Abnorm
al

Positive
(+)
Normal

Pea-sized fecal
material
2% Sodium
Carbonate

Analysis #29: Fecal Trypsin

Screens for: Cystic


Fibrosis and
Pancreatic
Insufficiency
X-ray filmstrip
immersed in diluted
fecal emulsion
Trypsin &
Chymotrypsin digest
gelatin on x-ray strip
(+) trypsin
complete digestion of
gelatin (blue tint)
(-) trypsin gelatin
Determination
remains intact

Cystic Fibrosis
Trypsin and chymotrypsin are
proteolytic enzymes. Their job is to
digest protein in the small intestine.
Normally, their precursors (their
inactive forms: trypsinogen and
chymotrypsinogen) are produced in
the pancreas and transported to the
small intestine.
In Cystic Fibrosis, mucous plugs can
block the pancreatic ducts that lead
into the small intestine, preventing
trypsinogen and
chymotrypsinogen from reaching
the intestine.
(-) trypsin in stool suggests Cystic
Fibrosis or pancreatic insufficiency

Analysis #30: Harrisons


Spot Test
(+) for
BILE!

Green =
biliverdin
Blue =
bilicyanin
Yellow =
choletelin

Tests for: liver function or blockage of bile duct


Filter: Barium Chloride + Urine --> add 2 drops of Fouchets reagent to
filter paper
Green = bilirubin is present

Analysis #31: Ehrlichs test


(Quantitative Test for
Urobilinogen)
Its NORMAL to
see a cherry red
ring
< 1:20

Cherry red ring in tubes >


1:20 means hemolytic
jaundice

Ehrlichs reagent
reacts with
urobilinogen to form
colored urobilinogenaldehyde
10% calcium chloride

Analysis #31: Ehrlichs test


(Quantitative Test for
Urobilinogen)
Check the tubes at an angle
No Cherry
Red =
NEGATIVE!

Cherry
Red =
POSITIVE
!

Negative in
the greatest
dilution
(1:320)

Slight
Positive in
tube 1:20

Analysis #32: Van Den Bergh


Reaction
(Qualitative Test for Bilirubin)

Violet Ring @
point of
contact =
(+) for
Direct
Bilirubin

If you
see

(+)
Urobilino
gen

(+)
Harrison
s

Partial Obstructive

If you
see

(-)
Urobilino
gen

(+)
Harrison
s

Complete Obstructive Jaundice

Example Practical Set Up


Jaundice

Qualitative

Harrisons
Spot Test

Normal

(+)
(<1:20)

(-)

(no jaundice)

Quantitative

Partial

(+)

(+)

Complete

(-)

(+)

Hemolytic

(+)
(>1:20)

(-)

Quantitative

Microscope Findings in
Feces

Vegetable Cells

Vegetable Cells

Vegetable Cells

Vegetable Fibers

Muscle Fibers

Muscle Fibers

Muscle Threads

Yeast Cells

RBC

Pus Cells

Fat Globules

Analysis #33: Blood


Grouping
Forward typing
determines antigens on
patient's or donor's cells
Cells are tested with the
antisera reagents anti-A,
anti-B

Reverse typing
determines antibodies in
patient's or donor's serum
or plasma

Anti-A
Sera

Anti-B
Sera

A w/ Agglutination

B w/ Agglutination

A w/ no agglutination

B w/ no agglutination

Forward Typing Table


Blood Type
Anti-A

(-)
(+)
(+)
(-)

Anti-B

(-)
(+)
(-)
(+)

O
AB
A
B

Reverse Typing
Known A Reagent
Cells

Known B Reagent
Cells

KA w/ Agglutination

KB w/ Agglutination

KA w/ no agglutination

KB w/ no agglutination

Reverse Typing Table


Known A

(-)
(+)
(+)
(-)

Known B

(-)
(+)
(-)
(+)

Blood Type

AB
O
B
A

Rh Typing

Agglutination = Rh(+)