CLINICAL PATHOLOGY

LIST OF INSTRUMENTS
[Link] E.S.R SET

[Link] SET(Wintrobe’s tube)

[Link]

4. HAEMOGLOBINOMETER
5. URINOMETER

6. ESBACH’S ALBINOMETER

7. STERNAL PUNCTURE NEEDLE

8. LUMBAR PUNCTURE NEEDLE

[Link] ESR SET
 Parts
1. Westergren stand

2. Westergren pipette
 Uses

[Link] of Erythrocyte
Sedimentation Rate(ESR)
PROCEDURE
 Procedure

Take 0.4 ml of 3.8 % sodium citrate

+
1.6 ml supplied blood
(1:4 Ratio)
❑ The fluid drawn up to mark of “0” in
Westergren pipette

❑ kept in straight position in Westergren

stand

❑ Reading after wait for one hour

followed by second hour.
Normal Values
1. Males : 0-5mm/ 1 Hour

2. Female : 0-7 mm/ 1 Hour

 What is ESR?

“The rate at which erythrocyte settle

down at the bottom of the pipette
at one hour is regards as
erythrocyte sedimentation rate”
 Stages of ESR
1. Stage of Rouleaux – 10Min

2. Stage of Sedimentation- 40 Min

3. Stage of Packing- 10 Min

 0ther Method

1. Wintrobe’s method
why ESR increased in chronic conditions
The serum contains both Cellular &
protein contents.

CELL Protein

If protein is increased causes red

cells to stick to each other(
Rouleaux Formation)
 In chronic condition ,there is high
proportion of Protein are present in the
blood leads to increased/fast Rouleaux
formation when compare to normal/slow
Rouleaux formation
 what anticoagulant used for ESR?
3.8 % sodium citrate

EDTA
2. HAEMOCRIT SET
(Wintrobe’s tube)
 Packed cell volume
• It is volume percentage
(%) of red blood cells in
blood
 Uses
1. To estimate of PCV(In the scale of
below upwards).

2. To estimate of ESR(In the scale of

above downwards).
ESR PCV
 Parts
1. Wintrobe’s tube

2. Wintrobe’s stand
PROCEDURE
1. The Wintrobe’s tube filled up with the
help of long capillary Pasteur’s pipette
and kept in stand for 1 hour for ESR
1st hour reading

2. Then it is centrifuged for ½ an hour at

3000 rpm and take the reading for
PCV
Normal Values
 [Link] POINTS Male Female
1. PCV(%) 39-49 33-43
[Link] Decrease in anemia.

[Link] Increase in polycythaemia

[Link]
 Parts
1. W.B.C. Pipette

2. R.B.C. Pipette

3. Improved Neubauer’s counting

chamber

[Link] Slip
 Uses
1. Total count of W.B.C in blood & CSF

2. Total count of R.B.C

3. Total count of platelets

4. Total count of spermatozoa

PROCEDURE
[Link] supplied blood up to 0.5 mark of
W.B.C pipette or R.B.C. Pipette.

2. Draw the W.B.C. fluid up to mark of 11 or

R.B.C fluid up to mark of 101 and mix
gently by rotation of the pipette in horizontal
position.

3. Discard 2 to 3 drops of solution from

the pipette and then put 1 drop of
diluted blood in between counting
chamber and cover slip.
4. Wait for 2 – 3 minutes

5. Adjust low power followed by high

power objectives.
i) W.B.C. - Count the 4 large square
(64 large Sq).

ii) R.B.C. - Count the 4 small square

in 4 corners and central
one square.(80 small sq)
Calculation
 Total count of W.B.C

• Total number of cells count in four

large square(64 Large sq) X 50

------------per cu mm of blood
 Total count of R.B.C

• Total number of R.B.C in five

small square(80 small sq) X 10000

---------per cu mm of blood
Normal Values
 Measurement Men Women
Hemoglobin (gm/dL) 13.6–17.2 12.0–15.0
Hematocrit (%) 39–49 33–43
Red cell count (106 /µL) 4.3–5.9 3.5–5.0
Reticulocyte count (%) 0.5–1.5
MCV 82–96(µm3 )
MCH 27–33 Pg
MCHC 33–37(gm/dL)
W.B.C (106 /µL) 4,000-11000
/cu mm
[Link]
 Parts
1. Haemoglobinometer tube
2. Haemoglobinometer Pipette
3. Comparator box
4. Glass stirrer
5. N/ 10 HCl Vial
 Uses

[Link] of total Hb%

PRINCIPLE
❖The blood is diluted with N/10 HCl
leads to formation of brownish-yellow
color (acid haematin hydrochloride)

❖The brownish-yellow color is matched

calorimetrically against the comparator
box by adding Distilled water (gm/100
ml)
PROCEDURE
- Hb tube filled with N/10 HCl up
to mark of 2.

- Draw the blood with the help of

Hb pipette (up to 20ul)

- Mix it with the glass stirrer

- Wait for 10 Minutes.

- Take Hb tube & Keep it in comparator
box and match with comparator column
after adding of Distilled Water.

- Read the lower level of fluid meniscus

from % scale or gram % scale directly
from Hb tube.
 Condition
1. Increased Hb - Polycythemia
Vera

2. Decreased Hb - Anemia
What is Haemoglobin
It is a conjugated protein
containing basic protein globin
(histone) united with four
prosthetic haem groups.
Normal Values
 Measurement Men Women
Hemoglobin (gm/dL) 13.6–17.2 12.0–15.0
[Link]
 Uses
For measurement of specific
gravity of urine sample.
 Parts
Glass tube – Marked by ½ , 1, 1 ½
& 2 oz. ( Below upward)

[Link] - Marked by
1000 – 1060 ( Above Downward)
 1000 1000
2 OZ
1010 1002
1004
1 1/2 1006
1020
1008

1010
1 1030

1040

1/2 1050

1060
PRINCIPLE
 “Principle of BUOYANCY”

Ability or tendency of something to

float in water or fluid
It is defined as “measure of the
weight of particles in solution”

Urea (20%), Nacl (25%), sulphate,

phosphate etc
 Specific Gravity

It is defined as the ratio between

the weight of a given volume of
urine and the distilled water at
room temperature
 Wt. of given volume of urine
S.G= Wt. of equal volume of D.W
PROCEDURE
1. The urine sample in glass tube
upto 1 ½ oz

a) if sample is hazy – centrifuge it

a) If the urine is not sufficient –add

distilled water.
2. Place the hydrometer inside the
glass tube.

3. Read of lower meniscus of the

graduation of hydrometer.
Result
1. If urine is sufficient then reading taken
directly.
2. If the urine is diluted then the last two digits
are multiplied with the diluting factor.
Example:
a) Reading of double diluted urine sample is
1010 will be calculated as,
10(first two digits) - 10(last two digits) X 2

S.G = 1020
Normal Values
Normal Specific Gravity- 1.010 – 1.030

❑Increased water intake- decreased S.G

❑Dehydration – increased S.G

 Increased Specific gravity

1. Diabetes mellitus

2. Nephrotic syndrome

3. Pyelonephritis

4. AGN
 Decreased Specific gravity

1. Diabetes incipidus

[Link] Glomerulonephritis
6. ESBACH’S ALBINOMETER
 Parts
1. Glass tube - Marked by “U’ and “ R’
and graduated from bottom to
upwards -1/2, 1, 2 to 12

2. Rubber cork

3. Wooden stand
 Uses

Quantitative analysis of total amount

of albumin (Percentage of albumin) in
gram per liter.
PRINCIPLE
“Principle of precipitation of albumin in the
presence of organic acids.
Organic acid – Negative ion
Protein- Positive ion

precipitation
PROCEDURE
❑Fill the urine sample upto ‘U”

❑Add Esbach’s reagent upto “R”

❑Gently mix up & closed by rubber cork.

❑Kept in dark place for 24hrs

❑Reading of yellow precipitate

Precaution
a) urine sample must be cleared by
Filtration

b) Urine must be acidic by adding acetic

acid.

c) The specific gravity of urine should be

in normal
Normal Values
 Normal value

Total urinary Excretion - <150mg/day

Normal Albumin < 30mg/day

Microalbuminuria 31-300 mg/day
Macroalbuminuria >300mg/day
 Protein serum Profile

Total Protein – 6.5 to 8.5 gm/dl

Albumin - 3.5 to 5.5 gm/dl

Globulin - 2 to 3.5 gm/dl

Alb: Glob - 1.5to 2.5

Albumin present in the urine is known as
albuminuria.
Condition:
1. AGN

2. Nephrotic syndrome

3. C.C.F

4. Acute Pyelonephritis
STERNAL PUNCTURE
NEEDLE
Type of Needles
Type of Needles
KLIMA Needle

SALAH Needle

JAMSHIDI Needle
SALAH Needle
 Parts
1. Needle proper

2. Guard

3. Stylet
 Site Of Puncture
1. Manubrium sterni or 1st or 2nd piece
of body of sternum

2. Posterior & anterior iliac crest

3. Medial aspect of upper end of tibia -

in case of children
 BONE MARROW 3000-4000ml

1500ml

LIQUID PART SOLID PART

RED BONE MARROW YELLOW BONE MARROW

( Hematopoietic stem cells) ( Mesenchymal stem cells)

RED BLOOD CELLS FAT CELLS

WHITE BLOOD CELLS

PLATELET CATILAGES
 BONE MARROW
ASPIRATION

With drawl of liquid portion of Bone marrow parts

BONE MARROW
Biopsy

With drawl of solid portion of Bone marrow parts

PROCEDURE
1. Antiseptic measure & 2 % Xylocaine
should be given as local anesthesia at
the site of puncture.

2. After fixing of guard, the needle

should be introduced along with stylet
by clock and anticlock wise direction
until feeling of sudden loss of
resistance.
3. About 0.2 ml marrow should be
aspirated and put on dry clean
smear slide and prepare a thick
smear and stain with Leishman
stain.
 Uses

To ensure that Bone marrow is

healthy to make normal blood
cells.
 Uses

Diagnostic purpose Therapeutic purpose

BM transplant
 INDICATION
1. Kala-azar

2. Aplastic anemia
 Complication

1. Injury to mediastinal structure

2. Haemorrhage
3. Shock
4. Infection
Normal finding
❑CELLULARITY →not less than 50%

❑ERYTHROID MATURATION → 20%

❑MEGAKARYOCYTES →15%
❑M:E RATIO → 2:1 - 4:1
❑WBC CELLS
promyelocyte – 2-4%
myelocyte- 10-18%
metamyelocyte -15-20%
segmented neutrophil- 15- 20%
lymphocyte -5-15%
Eosinophil- 1-3%
Plasma cells- 1-3%
Basophil- 0-1%
Monocyte- 1-2%
LUMBAR PUNCTURE NEEDLE

Spinal Tap
The recommended needle is a 22 gauge atraumatic needle. In our own practice we use a 22
gauge Whitacre needle, which is now the standard stock Lumbar Puncture needle in our
institution.
 Parts
1. Long needle about 8 cm long

2. Stylet
 USES

Collection Of Cerebrospinal
Fluid(CSF)
 Site Of Puncture

Intervertebral space between 3rd &

4th lumber vertebra
Birth – spinal cord extend upto L3

Adult – Spinal cord ends on inferior border of L1 or

superior border of L2
 Cerebrospinal fluid

It is the part of ECF

It is selective ultrafiltrate of plasma

Circulate Brain & Spinal cord in

subarachnoid space

Rate of formation-20ml/hr
(0.3-0.4ml/min)
 Function

[Link] (Buoyancy)

[Link]

[Link]
 Gross examination

Total quantity : 90-150ml/Adult

- 30-40ml – Ventricles
- 60-110ml- subarachnoid space
Colour- colorless & transparent
PH – 7.28-7.32 (Alkaline)
Appearance : clear, No clot formation
Cell count: 0-5 cells/cumm
Pressure: 90-180mm of H2O
 Chemical examination
Protein- 15-45mg/dl
Glucose – 45-80mg/dl
Sodium – 135-150mEq/L
Chloride – 120-130mEq/L

lactate – 10-25mg/dl (children-10-40mg/dl)

LDH – <2 - 7.2 U/ml

Others like urea, uric acid, lactic acid also present

Bilirubin is absent & NO RBC

PROCEDURE
 1. Position of the patient:

Patient should be kept on bed and

lying on one side of the body with
maximum bending of the spine to get the
maximum intervertebral space.
 Position

Lateral recumbent position with knees pulled up against

the chest.
2. Preparation of the site of puncture:

The site should be cleaned with

antiseptic and 2% of xylocaine
injected at the site of puncture as
local anesthetic.
 3. collection:

Collect 6-7ml of CSF fluid

between two vertebra(L3-L4) into the
space of the spinal cord.
 INDICATION

[Link] - spinal meningitis

[Link]
[Link] –arachnoid haemorrhage
[Link] haemorrhage
 CONTRAINDICATION

1. Local infection
2. Spinal cord tumor
3. hemorrhagic disorder
4. Septicemia
 Complication

1. Severe headache

2. Meningitis
Normal finding
 Cerebrospinal fluid Examination

❖ Appearance
❖ Cell count
❖ Gram stain
❖ Culture
❖ Protein determination
❖ Glucose determination
Points Normal Bacteria virus Fungus Tuberculosis

Appearance Clear Turbid Clear Slightly COBWEB

Colorless Large clot No clt turbid No clot
NO CLOT
WBC 0-5 >500 10-200 0-5 200-500
LYMPHO PMN LYMP LYMP LYMP
Pressure 90-180mm ↑↑ (>180) Normal/↑ ↑ ↑↑↑↑

protein 14-45mg/dl ↑↑ Normal/↑ Normal ↑↑↑↑

Glucose 50-80mg/dl ↓↓↓↓ Normal ↓ ↓

chloride 120- ↓ Normal ↓ ↓↓↓↓

130mEq/L