LABORATORY INVESTIGATION OF ANAEMIA

BY

PROF.YETUNDE. A. AKEN’OVA

Anaemia is an erythrocyte disorder characterized

by a haemoglobin concentration below normal for

the age and sex.

Anaemia can be categorized

1. A failure of production or shortened red cells survival- Intrinsic or extrinsic defect.

2. Anaemia of chronic disease consequent on chronic infection or inflammation or malignancy.

Haematocrit variation throughout life.Term newborn (cord blood) 44 %Term newborn (Capillary blood) 53 – 68Infant (3 months) 30 – 38Child (10 years) 37 – 44Pregnancy (30 wks gestation) 26 – 34 Adult female 37 – 47Adult male 42 – 54

Hct swings as high as 6-8% may occur with correction of dehydration of volume overload.

How was the blood specimen obtain Hgb/Hct values are frequently higher from a finger

stick than from a venous sample, unless excessive pressure is applied to facilitate blood flow.

Prolonged stasis from a tourniquet increases the haematocrit as do muscular activity and cold.

MCV = Hct RBC (Femtolitres)

MCH = Hgb RBC (Picograms)

MCHC = HgbHct ( g/dl RBC)

 

MCV – is a measure of cell size

MCH – is a measure of the average amount

of haemoglobin in each individual

cell.

MCHC – Is a measure of the concentration of

haemoglobin in each cell.

 Adult Male Adult Female

Hgb (g/di) 14 –18 12 – 16

Hct% 42 – 54 37 – 47

MCV (f1) 82 – 98 82 – 98

MCH (pg) 27 – 32 27 – 32

MCHC (g/dlRBCs) 31.5 – 36 31.5 – 36

PERIPHERAL SMEARCOMMON CAUSES OF VARIOUS RBC ABNORMALITIES

Hypochromia Iron deficiency Microcytosis Thalassaemia

Sideroblastic anaemiaChronic inflammatory

Macrocytosis Liver diseaseMegaloblastic anaemiaReticulocytosisMyelodysplastic anaemiaMyelophthysis

Marked Amsocytosis Marked iron deficiencyAnd Poikilocytosis megaloblastic

MicroanglopathicHaemolysisLeukoerythroblastosisHaemoglobinopathies

Target cells Liver diseaseHaemoglobinC,AC, SS, SCPost SplenectomyThalassaemia

Spiculated RBCs Hereditary AcanthocytosisLiver diseaseRenal disease

Post SplenectomyHypothyroidismMicroangiopathicHaemolysis

Tear drop cells LeukoerythroblastosisMegaloblastic anaemiaThalassaemiaErythroleukaemia

 

Howell – Jolly bodies Post Splenectomy

Megaloblastic Anaemia

Erythroleukemia

 

Spherocytes Hereditary Sphenocytosis

Auto immune haemolysis

Haemoglobin C disorders CC, SC.

Ovalocytes Hereditary ovalocytosis

Megaloblastic anaemia

Iron deficiency

Thalassaemia.

1.Cell size – Microcytic Macrocytic Normocytic

2. What is the basic Mechanism of the anaemia.a. Decreased affective marrow productionb. Bleedingc. Haemolysis

- Reticulocyte index – used to assess the appropriateness of the bone marrow

response to the anaemia.- The rate of Hct fall

Anaemia with appropriate reticulocyte response in the absence of bleeding means haemolysis.

ANAEMIAS WITH LOW MCV

Iron deficiencyThalassaemiaAnaemia of chronic diseaseSome haemoglobinopathiesSideroblastic anaemiasAluminium toxicity.

INVESTIGATION -     History and Physical examination -     complete blood count serum iron -     serum iron -     Total iron binding capacity -     Serum Ferritin -     Serum transferin receptor

SPECIAL INVESTIGATIONGIT –Radioraphic studies, Endoscopy, Gastroscopy, Esophagoscopy,Exploratory laparotomy

Blood film

Anaemia with high MCV >100flDifferential diagnosis1. Liver disease2. Alcoholism3. No associated disease4. Myelodysplastic syndromes5. Drugs6. Megaloblastic anaemias

Serum Vit B12

RBC folic acid Assay Bone marrow Aspiration

CAUSES OF VIT B12 DEFICIENCY IRON STAINDietary, Gastric, Terminal Ileum disease, fish tape worm.Pancreatic insufficiencyDrugsCongenital (transcobalamin 11 deficiency)

HAEMOLYSIS AND BLEEDINGAnaemia with a Normal or slightly elevated MCV and an appropriate Reticulocyte index.

Haemolysis would lead to Reticulocytosis Appropriate bone marrow response

An anaemia with an appropriate reticulocyte response in the absence of alert bleeding suggests haemolysis.

Retic count in a Normal Hct – 1%.

If haemolysis is chronic (months to years) the marrow production may reach a level of production greater than 10 times normal.Reticulocyte = Reticulocyte X Pt hct

Normal hct

ALLOANTIBODY VERSUS AUTOANTIBODY ALLOANTIBODY AUTOANTIBODY

Direct coombs’ Frequently negative. Positive

May be positive if

sensitized foreign

red cells are still

circulating.

Indirect Coombs’ Positive Positive or Negative

Antibody Screen

(Panel) specificity is seen Panagglutination no

specificity.

ANAEMIA WITH A NORMAL MCV AND LOW RETICULOCYTE INDEX

DIFFERENTIAL DIAGNOSIS

Renal failureAnaemia of inflammatory disease (Anaemia of Malignancy.etc)Mild (early) iron deficiencyCombined iron deficiency and megaloblastic anaemia Sideroblastic anaemiaAplastic anaemiaBone marrow infiltration (myelophthisis)Bleeding or hemolysis plus one of the above

Intravascular Haemolysis

When RBC destruction is rapid and occurs primarily Within

the vascular space, diagnosis is relatively easy.one sees the

following.

1. Haemoglobinaemia

2. Haemoglobinuria

3. Haptoglobin saturation

4. Haemosiderinuria

Extravascular haemolysis1. Autoimmune haemolysis2. Delayed haemolytic transfusion reactions3. Haemoglobinopathies4. Hereditary spherocytosis5. Hypersplenism6. Haemolysis with liver disease

The following cell types are seen1. Spherocytes2. Schistocytes3. Spiculated cells4. Bite cells

Low Haematocrit (Haemoglobin)

*Corrected Reticulocyte Count (Reticulocyte Index)

Normal value 0.5 – 2.5%

INVESTIGATE FOR:-Haemolytic Anaemias

(See Chart II)

INVESTIGATE FOR:-

Acute Blood LossEarly Iron DeficiencyInfectionInflammationNeoplasiaRenal Disease(Iron Folic Acid/B12)Sideroblastic AnaemiaRed cell AplasiaAplastic Anaemia

INVESTIGATE FOR:-

Folic Acid DeficiencyVitamin B12 DeficiencyLiver DiseaseBone Marrow FailureSideroblastic AnaemiaHypothyroidismChronic AlcoholismDrugs

Elevated

Low / Normal

Peripheral Blood FilmNormocytesMacrocytes

INVESTIGATE FOR:-Iron DeficiencyThalassaemiaSideroblastic AnaemiaAnaemia of Chronic Disease

Microcytes

Reticulocyte Index = (Reti culocyte Count (%) x Pt. Haemoglobin (g/dl)Expected Normal Haemoglobin (g/dl)

+ Correction Factor for Premature Release

+ Haemoglobin (g/dl)10 – 117 – 9<7

Correction Factor1.52.02.5

FLOW CHART FOR INVESTIGATING ANAEMIA

INTRACORPUSCULAR DISORDERS

Peripheral Blood Film

Osmotic Fragility

Acid Lysis (Ham’s Test)

Haemoglobin Electrophoresis

Red Cell Enzyme Assay

Family Studies

Normal

Anisocytosis, Poikilocytosis, Polychromasia, Nucleated Red

Blood Cells (NRBCs), Fragmented Cells

SHOWS

Heredity Spherocytosis

Paroxysmal Nocturnal Haemoglobin

Haemoglobinopathies (e.g. Hb S, Hb C)

Enzymopathies (e.g. G6PD and PK Deficiency)

Increased

Positive

Abnormal

Deficient

EXTRACORPUSCULAR DISORDERS

Peripheral Blood Film

Direct Antiglobulin Test

Clotting Studies

infection

Autoimmune Haemolytic Anaemia

Disseminated Intravascular Coagulation; Haemolytic Ureamic

Syndrome; Thrombotic Thrombocytopenic Purpura

Viral (e.g. CMV) Bacterial (e.g. Clostridium welchi)

Parasite (e.g. Malaria)

Positive

Prolonged

Normal

ASSESSING HAEMOLYTIC ANAEMIAS

Anaemia of chronic disease is characterized by1. Low serum iron concentration and defective

incorporation into haemoglobin despite adequate bone marrow stores of iron

2. A blunted erythropoietin response to anaemia. When mild it is normocytic normochromic but it can become severe and become hypochromic microcytic

3. Absoulte reticulocyte count is reduced, also ass.features indicative of chronic inflammation may be present.e.g. Neutrophilia, thrombocytosis, increased rouleaux formation.

Serum iron, serum transferin, are reduced while total iron binding capacity is increased.

Serum ferritin is increased, increase ESR,

plasma viscosity, reduced albumin and increased

fibrinogen 2 macroglobulin and - globulins are

increased