Lecture 3 - Erythrocyte Studies

Erythrocyte

Erythrocyte

carrier of hemoglobin for cellular respiration

• Manual hemocytometry

• Flow cytometry

• Electrical impedance

erythrocyte/discocyte/normocyte

counted by

BFU-E and CFU-E

erythrocytes are derived from

True

erythrocytes are not nucleated except for the immature forms (NRBCs)

Hemoglobin

a component of erythrocyte; red pigment; carrier of oxygen; respiratory pigment of RBC

• 1.39 ml Oxygen

• 3.47 mg Iron

1 gram of Hb =

2,3 diphospoglycerate

hemoglobin replaces ______________ in the heme portion

Oxygen is not sticking to the Hb

if the 2,3 diphosphoglycerate is low, it means that _________

Heme and Globin

composition of hemoglobin

4 pyrrole rings with iron

Heme is made up of

• 2 alpha, 2 beta chains (Hb A) = 94.5%

• 2 alpha, 2 delta chains (Hb A₂)= 3.5%

• 2 alpha, 2 gamma chains (Hb F) = 2%

Globin (with 2-3 DPG, 146 amino acids) composition

Hb A (94.5%)

most common globin

Hb F

predominates if Thalassemia

Myoglobin (140 amino acids)

respiratory pigment in skeletal and cardiac muscles

• Portland

• Gower I

• Gower II

3 types of Embryonic Hb

Abnormal Hb/ Hemoglobinopathies

• Hb S

• Hb BARTS

• Hb H

• Hb F

• Hb Lepore

• Hb M, Freiburg, Tubingen, St. Louis

• Hb Kansas, Caribbean, Hope, Seattle

• Hb Chesapeake, Koln, Gun Hill

• Hb C

• Hb Constant Spring

• Hb SC

• Hb E

Hb S

sickle cell anemia

common in African genetics

Hb BARTS

Alpha Thalassemia

Middle east & Africa

Hb H

Alpha Thalassemia

Hb H crystals stained with BCB

Hb F (in adults)

Beta Thalassemia (50% or greater)

Hb Lepore

Fusion of delta or gamma with beta chains

kind of beta thalassemia

• Hb M

• Hb Freiburg

• Hb Tubingen

• Hb St. Louis

HFTS

methemoglobinemia

• unstable Hb

• unoxidized

• heinz bodies

• BCB stain (brilliant cresyl blue)

• Wrights stain

• Hb Kansas

• Hb Caribbean

• Hb Hope

• Hb Seattle

KCHS

decreased oxygen affinity

(oxygen and hemoglobin only meet but don’t stick)

• Hb Chesapeake

• Hb Koln

• Hb Gun Hill

CKG

increased oxygen affinity

Hb C

Hb CC disease (Hb CC crystals)

Hb Constant Spring

few microcytic cells and target cells (type of Alpha Thalassemia)

Hb SC

Hb SC disease with few sickle cells and target cells

Hb SD

mild sickle cell anemia

Hb E

beta thalassemia (target cells)

True

anemic patients and newborns have high levels of Hb F

True

Hb F - cannot perform respiration

True

sickle cells don’t get affected by malaria

Brilliant Cresyl Blue

it stains methemoglobinemia

Sickle cells and Target cells

seen in sickle cell anemia

Sickle cells

drepanocytes and meniscocytes

Target cells

codocytes, planocytes, platycytes

• Hb SC

• Hb SD

the milder forms of Hb S

Abnormal Hb with no associated anemias

• Hb C - HARLEM ; Hb C- GEORGETOWN

• Hb D - PUNJAB

• Hb I

• Hb G-PHILADELPHIA

• Hb O - ARAB

these are what type of hemoglobin?

Porphobilinogen

seen in urine

Porphyrias

porphyrin is increased in urine

Porphyrias

anemia, heme is not producing, heme is deficient or decreased, indicates _________

• Oxyhemoglobin

• Deoxyhemoglobin/Carbaminohemoglobin

• Carboxyhemoglobin

• Sulfhemoglobin

• Methhemoglobin

types of hemoglobin

Oxyhemoglobin

found in capillary blood; bright red color

Carbaminohemoglobin / Deoxyhemoglobin

found in venous blood; dark reed

Carboxyhemoglobin

poisonous; cherry red color

Sulfhemoglobin

lavender color

Methemoglobin

chocolate brown color

Shift to the RIGHT

decreased affinity of Hb for O₂

Shift to the LEFT

increased affinity of Hb for O₂

Shift to the RIGHT

• Increased temperature

• Increased 2,3-DPG

• Increased CO₂

Shift to the LEFT

• Increased pH

• Increased Hb F

1. Direct Visual

2. Acid Hematin

3. Alkali Denaturation Test

4. Spectrophotometric

5. Gravimetric (Copper Sulfate)

6. Acid Elution Slide Test

methods for hemoglobin determination

Direct Visual

• oldest method for determining hemoglobin

• qualitative method

• Tallquist, DARE

Acid Hematin (hemometer)

• utilizes acid to lyse RBC to release hemoglobin

• semi-quantitative

• Tests

  • sahli-hellige

  • sahli-hayden

  • sahli-adams

  • hayden-hausser

  • newcomer

  • osgood-haskin

2%

normal percent of Hb F

WU, CLEGG, KING

type of test in Alkali Denaturation Test

Spectrophotometric

• gold standard for hemoglobin

• Drabkin’s reagent (requires standard)

darker color = high Hb conc

lighter color = lower Hb conc

Gravimetric (copper sulfate)

used by Philippines Red Cross to test for the specific gravity of the capillary blood

1.053

specific gravity =

not okay to donate

Float

okay to donate

Sink/Lower

Acid Elution Slide Test and Alkali Denaturation Test

Hb F

Citric acid phosphate

remove the Hb

Hb F —> Thalassemia

a lot of bright pink

Male: 13.5 - 17.5 g/dL

Female: 12-16 g/dL

reference values for Hb (not SI unit)

Male: 135-175 g/L

Female: 120-160 g/L

reference values for Hb (SI unit)

1. Adam’s Micromethod

2. Wintrobe / Macromethod

methods for Hematocrit / Packed Cell Volume determination

Adam’s micromethod

• capillary tube

• centrifuge (10k-13k rpm for 5 mins)

• microhematrocrit reader

Wintrobe method / Macromethod

• wintrobe tube

• centrifuge (3k rpm for 15 mins)

Hct % = height of packed cell / (height of packed cell + height of plasma) x 100

formula for Hct

Male: 0.41 - 0.53 liter/liter

Female: 0.36 - 0.46 liter/liter

reference values for Hct (SI unit)

Male: 41-53%

Female: 36-46%

reference values for Hct (percentage)