Lab 5 RBC Flashcards by Lea Damler

What will happen when there is a problem with RBCs?

Tissue hypoxia

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What are the most important parameters for detecting pathological processes?

RBC count

Haemoglobin concentration/function

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How can Hgb be measured?

Drabkin method (Spectrophotometry)
Potassium ferricyanide added to whole blood
RBCs hemolyzed, Fe3+ is formed from Fe2+
Cyanmethemoglobin is the orange coloured end product, measired at 540 nm

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What is normal Hgb content in the blood?

18-20 mmol/l or 120-180 g/l

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In case of intravascular hemolysis, what change in Hgb conc is expected?

No change

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Upward shift in oxygen dissociation curve

Oxygen binding capacity increased by:
hypothermia
low 2-3 DPG conc (stimulates O2 release)
alkalosis
CO (cherry colour)

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Downward shift in oxygen dissociation curve

Oxygen binding capacity decreased by:
decreased affinity
hyperthermia
high 2-3 DPG conc
acidosis

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2,3-DPG

Diphosphoglycerate
Prod by RBC during glycolysis
Promotes release of O2, binds to deoxygenated Hgb near respiring tissue to ensure tissues that need O2 get it

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SAT %

Oxygen saturation
The percentage of oxygenated Hgb molecules compared to the whole amount of Hgb molecules in one unit of blood
Arterial: 95-99%
Venous: 80-90%

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Methemoglobin

Hemoglobin containing Fe3+ (ferric) oxidized form
Cannot take up oxygen
Reduced back to Hgb by methemoglobin-reductase enzyme

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What can cause severe oxidative damage to RBCs?

Nitrites, free radicals, paracetamol, onion

Leads to methemoglobinaemia

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What are some symptoms of methemoglobinaemia?

Dark brown colour of blood

Deeply cyanotic mucous membranes

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Who has Hgb that are most sensitive to oxidative damage?

Cats and newborns of any species

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How can you treat severe methemoglobinaemia?

Blood transfusions and antioxidants

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How can we give a rough estimation of Hgb?

Suspecting normal mean Hgb conc of RBC (MCHC)

PCV / 3 x 1000 = Hgb g/l

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Give some causes of increased Hgb concentration

relative polycythaemia: dehydration

absolute polycythaemia: RBC leukemia, high altitude, greyhound, llama

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Give some causes of decreased Hgb concentration

relative oligocythaemia: hyperhydration

absolute oligocythaemia: anaemia

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What species have Hgb conc affected by age?

Swine

Younger have lower Hgb conc than older swine

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Give some reasons for intravascular hemolysis

DIC
Thrombocytopenic purpura
Hemolytic uremic syndrome
Mechanical stress

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Give some reasons for extravascular hemolysis

Hypersplenism

Inherited: sickle cell anemia, spherocytosis

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Give methods to measure RBC count

Bürker chamber (lol)
Estimation
Automatic cell counter

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What sample is used for Bürker chamber and what dilution?

EDTA whole blood

200x dilution

How is RBC count estimated?

If we suspect a normal average RBC volume (MCV)

Ht / 5 x 100 = RBC count x 10^12/l

RBC count by automatic cell counter

Size dependent: electric impedance
40-100 fl
histogram is provided from the results

What can we expect to see on the ACC histogram during immunohemolytic anaemia?

RBCs form aggregates, they are not counted

Normal RBC count

4.5-8 x 10^12/l (teraliter)

Histogram expected during non-regenerative anemia

Low RBC count | narrow RDW

Histogram expected during regenerative anemia

Low RBC count | BUT wide RDW and larger cells (reticulocytes)

Histogram expected during microcytic anemia

Low RBC count | "left shift" small cells

Histogram expected during macrocytic anemia

Low RBC count | "right shift" larger cells

MCH

``` Mean corpuscular hemoglobin Average Hgb content of RBCs HGB/RBC count Normal: 20 pg PER CELL (but young can be higher) Hypo/hyperchromasia ```

MCV

``` Mean corpuscular volume Indicates average size of RBCs PCV/RBC count Normal: 60-70 fl Micro/macrocythaemia ```

What species and ages do you see differences in size of RBCs?

``` Fe/Eq smaller RBCs Young RBCs are bigger Newborns have bigger RBCs Akita inu: small RBCs Poodle: large RBCs ```

Causes of microcytosis

Chronic blood loss Fe, Cu, B6 deficiency Portosystemic shunt

Causes of macrocytosis

Polycythaemia absoluta vera | B12, Cobalt, folic acid deficiency

MCHC

Gives a ratio: sees if the RBCs are large or small too MCH/MCV Normal: 300-350 g/l Hypo/hyperchromic

Decreased MCHC

Hypochromasia Newborns Regenerative anaemias Iron deficiency anaemias (also Cu/B6)

Increased MCHC

``` Hyperchromasia Acute erythroleukemia (polycythaemia vera) B12, folic acid and cobalt deficiency Immunohemolytic anaemia (spherocytosis) Lead poisoning Splenectomy ```

What changes in derived parameters are seen during regenerative anaemias?

Increased MCV, decreased MCHC | Macrocytosis, hypochromic (inc reticulocytes)

What changes in derived parameters are seen during non-regenerative anaemias?

Normocytic, normochromic | Normal or decreased MCH

What changes in derived parameters are seen during non-Fe/Cu/B6 deficiency anaemias, liver failure or portosystemic shunt?

Microcytic, hypochromic | Decreased Hgb synthesis

What is pyridoxine needed for in blood production?

Porphyrine production (Hgb)

What is copper needed for in blood production

Iron oxidation (ceruloplasmin)

What changes in derived parameters are seen in Akita inus?

Microcytic normochromic

What changes in derived parameters are seen during | FeLV infection, B12, Co, Folic acid deficiency, erythroleukemia and poodle macrocytosis?

Macrocytic normochromic

RDW

Red cell Distribution Width Range of average size of RBCs Derived from the histogram at 20% of the peak High RDW = high variation in size (positive) Short = non-regenerative process

PDW

Platelet Distribution Width

How can you differentiate RBCs from reticulocytes in a blood smear?

Basophil punctuates stained by brylliant-cresil blue stain The punctuates are rRNA remnants Large punctuates: young reticulocyte

Species differences reticulocytes

Cats: punctuated form more common (older) | Eq/Ru: no reticulocytes are found in blood, only in bone marrow

What are nucleated RBCs called?

Normoblast | Cannot carry blood

Can reticulocytes perform the same tasks as RBCs?

Yes, they can carry oxygen

What blood sample and what stain is used for reticulocyte count?

EDTA fresh whole blood Brylliant-cresil stain The cells have to be alive: 2-3 hrs needed in room temp for the reticulocytes to take up the stain

When and why should the reticulocyte count/percentage be corrected?

When the animal is sick Correlated to the RBC count or the PCV Because mature RBCs are more sensitive to damage than young RBCs and reticulocytes