ESR, PCV AND BLOOD INDICES

DR. JANANI MATHIALAGAN1ST YEAR POSTGRADUATE PATHOLOGY

OBJECTIVES OF THIS CLASSTo perform and interpret ESRTo perform and interpret PCVTo perform and interpret Blood Indices

ERYTHROCYTE SEDIMENTATION RATE (ESR)

ESR is the measurement of the rate of sedimentation of red cells in anti-coagulated blood.

Blood is allowed to stand for 1 hr in an open-ended glass tube mounted vertically on a stand

Length of column of plasma above the red cells is measured in mm.

Anticoagulated blood is drawn up into a tube of standardized dimensions and left in a vertical position for exactly one hourBy that time, the red cells would have separated and settled from the plasma.Upper plasma column is recorded by reading from the scale on the side of the tube.Measures the distance that RBCs will fall in a vertical tube over a given time periodInitial screening tool and also as a follow-up test – monitor therapy and progression or remission of disease

Three definite phases:

• First or Lag Phase (10mins) – red cells form a characteristic rouleaux pattern (aggregation) and sedimentation is generally slow. (Pack of coins)

• Decantation Phase (40mins) – The rate accelerates in this phase; fast settling or sinking of RBCs

• Final Packing Phase (last 10mins) – slows again as red cell aggregates pile up at the base of the tube. There is slow sedimentation.

FACTORS AFFECTING ESR

1. Plasma factor2. RBC factor3. Technical factor

PLASMA FACTORSIncreased fibrinogen increases rouleaux formation thereby increasing ESR

S. haptoglobulin , C - reactive protein & cholesterol also increases ESR

Albumin and lecithin decreases sedimentation i.e. decreasing ESR

RBC FACTORS

Primarily through changes in number and/or shape

Anemia responsible for increased ESR◦ Microcytes – sediment more slowly◦ Macrocytes – sediment faster

Poikilocytosis retards ESR because abnormal shape hampers rouleaux formation

Anticoagulants – Sodium citrate & EDTA doesnot affect ESR

oxalates & heparin may affect

TECHNICAL FACTORS

Poor temperature controlLength and bore of the tubeVibrationVerticality

METHODSWestergren’s method

Wintrobe’s method

Micro - ESR

Automated systems

Zeta sedimentation

The method for measuring the ESR recommended by the International Council for Standardization in Haematology (ICSH)

Based on that of Westergren, who developed the test in 1921 for studying patients with pulmonary tuberculosis.

CONVENTIONAL WESTERGREN METHOD

The recommended tube is a straight glass or rigid transparent plastic tube 30 cm in length2.55 mm in diameter. Bore must be uniformA scale graduated in mm extends over the lower 20 cm.

For the diluent, 3.8 g/dl Trisodium citrate used

Dilution – 1:4 0.25ml trisodium citrate : 1ml blood

Mix the blood sample thoroughly and then draw it up into the Westergren tube to the 200 mm mark by means of a rubber teat or a mechanical device

Place the tube exactly vertical and leave undisturbed for exactly 60 min, free from vibrations and draughts and not exposed to direct sunlight.

Then read to the nearest 1 mm the height of the clear plasma above the upper limit of the column of sedimenting cells.

Westergren pipette filled with blood and placed vertically on the rubber cork in the rack

ERYTHROCYTE SEDIMENTATION RATE

Average ESR value by Westergren Method:Male – 3-5mmFemale – 4-7mm

PROCEDURE – WINTROBE METHOD 1. Add well mixed double oxalate / EDTA blood to the zero mark of the Wintrobe tube, using a pipetteAvoid air bubbles

2. Place in vertical position in a rack and let sit for 60 minutes 3. Read and record results in millimeter (distance which the cells have settled)

Average ESR value by Wintrobe’s Method:◦ Males: 0 – 9mm/hr◦ Females: 0 – 20mm/hr◦ Children: 0 – 13mm/hr

Increased ESR

•Chronic infections e.g. Tuberculosis•Extensive/ Chronic inflammation•Collagen vascular disorderso Systemic Lupus Erythromatosuso Rheumatoid asthritiso Systemic Sclerosis•Shock•Active syphilis•Active infectious infections

Decreased ESR

•Newborns•Congestive heart failure•Polycythemia•Marked leukocytosis•Allergic states•Sickle cell anemia

Ratio of volume of RBCs to that of whole blood It indicates relative proportion of red cells to plasmaExpressed in percentage.Also called hematocrit or erythrocyte volume fraction

PACKED CELL VOLUME

Methods:1. Macrohematocrit method (Wintrobe Method)2. Microhematocrit method3. Electronic Method

WINTROBES METHOD•Wintrobe’s tube – 110mm long,

internal bore 2.5mm &

a flat inner base.

Graded 0-10 on both sides.

Method: 1.Mix the anticoagulant blood sample thoroughly2.Draw blood in a Pasteur pipette3.Fill the tube upto 10 mark4.Centrifuge the sample at 2000-2300 rpm for 30mins

5.Take the reading of the length of the column of red cells

Buffy coat- WBC& PLATELETS.

UPPER MOST LAYER – PLASMA

•Yellowish-Jaundice

•Pink-haemolysis

•Milky-hyperlipidemia

PCV reading

PRECAUTIONS Use recommended amount of EDTA

Test done with in 6-8 hours

Wintrobe tube should be filled from below upwards so that no air bubble is trapped.

INCREASED PCV

Polycythemia-Newborns, High altitude, Hypoxia due to lung and heart diseases.Congestive Heart failure, Burns (loss of plasma), Dehydration, Severe Exercise, Emotional stress

DECREASED PCV

Anaemia

Pregnancy (Hemodilution)

RED BLOOD CELL INDICES:RED BLOOD CELL INDICES:

1.Mean corpuscular volume(MCV)2.Mean corpuscular hemoglobin(MCH)3.Mean corpuscular hemoglobin concentration(MCHC)4.Red cell distribution width (RDW)

Mean Corpuscular Volume(MCV)

Average or mean volume of a single red blood cell Expressed in femto liter (fl)

Calculation Formula:

MCV = PCV in percentage X 10

RBC count per cmm

Normal range:

80-100 fl

INCREASED

Megaloblastic anaemiaChronic alcoholismLiver diseasenewborns

DECREASED

Microcytic hypochromic anaemia

Mean Corpuscular Hemoglobin(MCH)

Average hemoglobin content (weight of Hb) in a single red blood cell Expressed in picograms(pg).

Calculation:Formula:

MCH = Hb in gm/dl X 10 RBC count per cmm

Normal range: 27 – 32 pg

INCREASED

Macrocytic anaemiaNewborns

DECREASED

Microcytic anaemia

Mean Corpuscular Hemoglobin Concentration(MCHC)

Concentration of haemoglobin in 1 dl or 1 liter of packed red cells

Calculation Formula:

MCHC = Hb in gm/dl X 100 PCV in %

Normal range: 30 – 35 g/dl

INCREASED

Hereditary spherocytosis

DECREASED

Hypochromic anaemia

CLINICAL SIGNIFICANCE A) Macrocytic anaemia:

◦ MCV slightly increased upto 150 fl◦ MCH is slightly increased◦ MCHC is normal or diminished

B) Microcytic anaemia:◦ MCV is diminished up to 50 fl or lower◦ MCH is diminished to 15 pg or lower◦ MCHC is diminished to 20% or less

C) Spherocytosis:◦ MCV is diminished◦ MCHC is elevated

RED CELL DISTRIBUTION WIDTH

Measures the degree of variation of red cell size in a blood sample.

Increased in iron deficiency anaemia Decreased in beta- thalassemia trait

Normal value: 9.0-14.5