Thalassemia and Sickle Cell Hemoglobinopathy


Embryonic Hgb


2 Zeta Chains &2 Beta, 2 Gamma, or 2 Delta Chains


Fetal Hgb


2 Alpha Chains2 Gamma Chains


Adult Hgb A1


2 Alpha Chains2 Beta Chains


Adult Hgb A2


2 Alpha Chains2 Delta Chains


Normal Hgb Distribution


Hgb A1 96%Hgb A2 2-3%


Abnormal Hgb Fractions


Hgb SHgb CHgb HHgb EHgb MBart's Hgb


Abnormal Structures due to?


Amino acid substitutions on globin chains.


Types of abnormal structures.


Sickle Cell AnemiaSickle Cell TraitHgb C DiseaseHgb S-C Disease


Abnormal Synthesis


Due to manufactor of Alpha or Beta ChainsAlpha ThalassemiaBeta Thalassemia


Hgb S


#6 Amino Acid on beta chains is replaced with Valine.Inherited as Autosomal recessive gene.Homozygous is SSHeterozygous is AS


Hgb C


#6 Amino Acids on beta chains replaced with lysine.Inherited with Autosomal recessive gene.Homozygous state 93-100% Hgb CCommonly inherited in combination with Hgb S


Hgb E


Lysine substituted for #26 amino acid on beta chains.Autosomal recessive.Homozygous Hgb E is microcytic anemias with target cells.Heterozygous is silent carrier.30% of southeast asians have Hgb E.


Hgb M


Tyrosine is substituted for histadine on either Beta or Alpha Chains.Hgb M is methomoglobin (Fe + 3)Autosomal DominantPatient Appears CyanoticBlood Appears Brown.Heinz Bodies Present.


Hgb H


4 beta chains


Bart's Hgb


Globin structure is 4 gamma chains.


Hereditary Persistance of fetal Hgb (HPFH)


Continued production of Hgb F into adulthood.Homozygous 100% Hgb FHeterozygous 30% Hgb F


Detection of Hgb S


Abnormal RBC Morphology on smear.Positive screening method.Confirm with electrophoresis.


Sickle Cell Anemia


Normocytic/ Normochromic AnemiasSickle CellsTarget cellsNRBCsHowell- Jolly BodiesPolychromatophilia (increased reticulocyte)


Screening Hgb S


Cappillary StasisMetabisulfateDithionite (Sodium Hydrosulfate)


Metabisulfate Method


Hgb is insoluble in low O2 concentration & perpetuates out.Metabisulfate chemically lowers concentration of O2 in the sample causing sickling of the cells.


Metabisulfate reagents


EDTA Whole Blood2% metabisulfate


Metabisulfate Method


Place 1 drop of EDTA whole blood on slide.Add 2 drops 2% metabisulfate to blood and mix.Coverslip and seal with vaseline.Let stand for 30 minutes.Observe for Sickling.


Dithionite Method Principle


Hgb S insoluble in low O2 concentration.Dithionite chemically lowers O2 Concentration in test sample causing Hgb to PPT out.Precipitation of Hgb S causes turbidity.


Dithionite Method Reagents


EDTA Whole BloodSaponin (Lysing Agent)Dithionite (sodium hydrosulfate) ASA reducing agent.


Dithionite Method Procedure


Add Saponin and Dithionite reagent to a test tube.Add blood sample to reagent & mix.Allow sample to sit at room temp for 10-15 minutes.Appearance of turbidity is positive for Hgb S & C.


Hemoglobin Electrophoresis


Movement of charged particles in an electric field.Negative charged particles migrate toward positive pole (anode)Alkaline Ph gives proteins a negative charge.


Types of Hgb Electrophoresis


Cellulose Acetate @ Alkaline Ph.Citrate Agar @ Acid Ph


Electrophoresis Cellulose Acetate Order


Hgb A1 closest to anode.F/G nextS/D nextA2Hgb C/E closest to cathode.


Electrophoresis Citrate Agar Order


Hgb F closest to anodeThen A1,G,D,ES nextHgb C closest to cathode & behind application point.


Thalassemias


Due to Abnormal synthesis of Hgb chains.Most common inherited disorder worldwide.


What type of cells doThalassemias produce?


Produce Microcytic Hypochromic Anemia with poikilocytosis.Schistocytes, NRBCs, target cells, basophilic stipling.


Types of Thalassemia


Alpha Thalassemia- production of alpha chains is affected.Beta Thalassemia-Production of beta chains is affected.


Alpha Thalasemia


Production of Alpha chains is abnormal.All hemoglobins affected.


Distribution of Alpha Thalassemia.


Middle East, Orient, SE Asia25% of Thailand have Alpha Thalassemia.30% of African Americans are silent carriers.


Distribution of Beta Thalassemia.


Production of beta chains is abnormal.Seen in Mediterranean Populations.Jewish, Greek, Italians.5-10% of population are heterozygous.


Autosomal Dominant Minor.


Heterozygous StateAlpha Thalassemia MinorBeta Thalassemia Minor


Autosomal Dominant Major


Alpha Thalassemia MajorBeta Thalassemia Major.


Beta Thalassemia Inheritance


2 Genes for productions of beta chainson CH#11Called B0 & B+B0 produces no beta chains.B+ has decreased productions


4 Patterns of Beta Thalassemia Inheritance


Silent CarrierB Thalassemia MinorB Thalassemia MajorThalassemia Intermedia


Beta Silent Carrier


Production of Beta chains is nearly normal, slightly decreased.No Hematological Abnormalities noted.


Minor Beta Heterozygous


Mild microcytic hypochromic anemia.Target cells, Elliptocytes, & basophillic stippling seen on peripheral smear.Conc of Hgb A2 & Hgb F are increased.


Beta Thalassemia Intermedia


Minimum Hgb of 7g/dl w/o transfusion.NRBCs seen on peripheral smear.Concentration of Hgb A2 & Hgb F are increased.


Alpha Thalassemia Inheritance


4 genes for the production of alpha chains on CH#16.Inherit 1 from each parent Alpha 1 & Alpha 2.


Alpha Thalassemia gene defects


No production = no functioning gene on CH#16Decreased Production= 1 functioning gene on ch#16


4 Inheritance of Alpha Thalassemia.


Silent CarrierAlpha Thalassemia Minor (Heterozygous)Hgb H disease.Alpha Thalassemia Major (Hydrops Fetalis)


Silent Alpha Carrier


Due to deletion of 1 gene with 3 functioning genes.Production of Alpha Chains is affected only slightly.Hematologically normal appearance.


Alpha Thalassemia Minor


Heterozygous.Due to only 2 functioning genes.Individual produces 2-10% Burt's Hgb.Bart's Hgb is 4 gamma strains.Hematologically mild microcytic anemia.


Hgb H disease.


Only 1 functioning gene.Common in southeast Asia.


Alpha Thalassemia Major


Hydrops FetalisNo Functioning genes.Produced 80% bart's HgbNo functioning ACauses death in utero or shortly after birth.


Bart's Hgb Alpha Thalassemia association.


1 gene deleted - 1-3% bart's2 genes deleted 4-10% bart's3 genes deleted Hgb H disease 15-25% bart's & 10-25% H4 genes deleted Fetal Death 100% bart's