Clinical Haemotology 1

Clinical Haemotology 1

• RBC Production and Structure

ILOs

1. Understand the concepts behind the laboratory assessment of the erythrocyte
2. Recognise common RBC abnormlaities, including terminology and clinical significance
3. Be aware of the other auxiliary diagnostic tests used in the assessment of RBC abnormalities

RBC Production and Structure

Production

• Haemotology - the study of blood and the body tissues that make it
• Erythropoiesis - the production of erythrocytes:
• Mainly in bone marrow - medullary cavity of long bones
• In spleen and liver - if bone marrow fails, times of high demand
• Stimulated by erythropoietin (EPO) production

Structure

• Biconave disks - allows increased SA for oxygen diffusion
• Flexible - pass through small vessels
• Cytoplasm contains haemoglobin
• Anucleate = unable to regenerate
• Senescent RBC - end of life EBC

EPO

70

110

145

160

1-2 days

5-7 days

Understand the concepts behind the laboratory assessment of the erythrocyte

PCV

Manual

Blood Smear

Manual

Haemotology

Analyser

Haemotology

WBC Parameters

• WBCs
• Neutrophils
• Lymphocytes
• Monocytes
• Eosinophils
• Basophils

Tube Types

Buffy Coat

Plt Parameters

• Platelets

RBC Parameters

• Haemoglobin
• Haematocrit
• RBC
• Meal Cell Volume
• Mean Cell Hgb
• Mean Cell Hgb conc

Bold = measured by analyser Italic = calculated

Artefacts

Mass

Erythrocytes

Indices

PCV

Interpretation

• Packed Cell Volume - RBC mass measurement
• Different to Hct - which is calculated
• Findings:

• % of RBC in a volume of blood
• Buffy coat assessment [WBC and Plt}
• Plasma colour assessment
• Total protein measurement

• High - erythrocytosis:
• Physiological - breed
• Pathological - dehydrated, splenic contraction, increased EPO
• Artefact
• Low - erythropenia:
• Pathological - IVFT overdose, anaemia
• Artefact

PCV and TS

Method

Collect Sample

EDTA

Fill capillary tubes

2 for balance

Invert Gently

Stop 3/4 full

Plug tubes

Clay

Centrifuge

10,000 rpm, 5 mins

Read PCV

Normal PCV

Blood Smear

Normal RBC

• Three main parts to the blood smear:
• Base
• Monolayer
• Feathered edge
• Method:
• Drop blood on slide
• USe second slide to create smear
• Stain smear and use oil to visualise under microscope
• Begin with feathered edge:
• Platelet clumps
• Atypical cells
• Review monolayer - battlement style:
• RBC morphology
• Platelet counts
• Review the lateral edge:
• WBC morphology and counts

Recognise common RBC abnormlaities, including terminology and clinical significance

Schistocytes

Acanthocytes

Macrocytes

Size

Shape

Codocytes

Anisocytosis

Poikilocytosis

Echinocytes

Microcytes

Elliptocytes

Keratocytes

Abnormalities

Spherocytes

Agglutination

Hypochromasia

Colour

Pattern

Polychromasia

Reticulocytes

Rouleaux

Polychromatophils

Ghost Cells

Inclusions

Basophiliic Cytoplasm

Howell-Jolly Bodies

Siderocytes

Heinz Bodies

Be aware of the other auxiliary diagnostic tests used in the assessment of RBC abnormalities

Clinical Haematology Task Review Seminar - Bill

• Ddx for pallor:
• Anaemia
• Hypoperfusion - hypovolaemia, CVD, iatrogenic [alpha2 agonist]
• Stress
• Regenerative anaemia causes - haemorrhage, haemolysis
• Non-regenerative anaemia causes - BM dz, Fe deficiency
• Bone marrow needs iron to make hemoglobin - without enough iron, the body can't make enough hemoglobin for red blood cells
• Haemolysis causes - divided in IM processes, infectious dz
• IMHA
• Shear injury
• Angiostrongylus
• Ox damage - paracetamol, onions, garlic
• Rodenticide
• FIP
• Haemotropic mycoplasmas - obligate parasites on RBC surface

Clinical Haematology Task Review Seminar - Luna

• Leukogram - inflammatory [not stressed]:
• Presence of band neutrophils indicative
• Go to seminar slides for left shift explanation
• Left shift - usually means infection but not always:
• Could be inflammatory
• Also slides on pancreatitis

Clinical Haematology Task Review Seminar

• EDTA blood sample left in heat for a long time - haemolysis
• Clotting cascade activated as soon as BV is ruptured - by needle for sample
• If haemolysis has occured - plasma will be tinged red and MCHC increased
• Mean corpuscular hemoglobin concentration (MCHC) refers to the average concentration of hemoglobin inside a group of red blood cells
• Goes up because there is the same amount of Hgb present [in cells and now lysed in plasma] but there are less cells [due to lyse] so conc. cf to RBC count increased
• Lipaemic - Hgb won't dissolve, and will only dilute in plasma
• Changes that would be seen in EDTA sample 3 days post collect:
• HCT increased [as RBC swells; HCT assess RBC mass]
• MCV increased [as RBC swell]
• MCHC reduced [as volume increases, conc will decrease; Hgb amount is fixed but if RBC swell than the Hgb amount is now in larger volume so becomes diluted = less concentrated]
• Mass is the amount of matter an object contains, while volume is how much space it takes up
• Haemolysis occurs >3 days post storage in EDTA - resulting in hyperkalaemia [K in cell now released and is in blood] [Na higher in plasma]

• RBC that have ruptured in circulation and lost Hgb
• NB as Hgb in circulation can cause AKD
• RBC membrane remains - ghost
• Representation of haemolysis
• Causes:
• Oxidative injury - onions and garlic
• Infectious dz - clostridium, lepto
• Hypophosphataemia - decreases ATP levels and increases haemolysis susceptibiliy
• Lipaemic animals [not fasted]
• Artefact

• Large RBC
• Hgb content - normal
• Causes:
• Regeneration [when + polychromasia] - younger RBC
• Abnormal DNA synthesis:
• FeLV - virus enters RBC and prevents DNA metabolism
• Familial - toy poodle
• Laboratory artefact

Diagrams

• ​Iron-containing metalloprotein
• Carries O2
• Breaksdown into haem and globins
• Globins - broken down to aa
• Haem breakdown produces unconjugated bilirubin:
• Unconjugated = fat soluble and unable to be excreted
• Taken up by albumin to be transported to liver
• 20% dissociates and enters liver
• Bilirubin in liver taken up by hepatocytes
• Bilirubin conjugated to glucuronic acid = now water soluble

• Bile production:

• Conjugated bilirubin contributes to bile production [as well as cholesterol, phospholipids and bile salt]
• Bile - flows through hepatic duct into gallbladder [except horses]
• Gallbladder conc and stores bile
• Bile secreted into small intestine via common bile duct
• Conjugated bilirubin in bile not absorbed in duodenum or jejunum due to water-soluble properties:
• Promotes excretion of bilirubin via faeces
• Some conjugated bilirubin is deconjugated by mircoflora in ileum and colon:
• Majority on unconjugateed bilirubin returns to liver via enterohepatic circulation [recycled as heme?]
• Rest excreted via faeces

Jaundice

• Measures RBC mass [same as PCV]:
• High = erythrocytosis:
• Dehydrated
• Splenic contractions
• Breed [greyhound]
• Artifact

• Low = erythropenia:
• Anaemia
• Overdilution
• Artifact

• Reticulocytes - immature RBC that contain RNA:
• Larger and basophilic due to RNA
• Normal physiology:
• Remain in BM for 2 days
• Released and lose RNA - done by spleen
• Mature to RBC

• Polychromatophils - immature reticulocytes:
• Contain more RNA than reticulocytes
• All polychromatophils = reticulocytes
• All reticulocytes ≠ polychromatophils

• If anaimal is anaemic - # polychromatphils indicates if BM is responding:
• Many polychromatophils = regenerative anaemia