Sickle Cell and ESR laboratory methods.ppt

Sickle Cell and ESR laboratory methods.ppt

• 1.
  Sickle Cell Sickle Cell 1
• 2.
  Genetics Review Genetics Review Chromosomes Chromosomes Genes Genes Loci Loci Genotype Genotype 2
• 3.
  Normal Adult Hemoglobin(Hgb A) Normal Adult Hemoglobin (Hgb A)  Heme molecule composed of iron in the Heme molecule composed of iron in the ferrous state (Fe ferrous state (Fe++ ++ ) )  Globin is the protein portion Globin is the protein portion  95-98% of adult Hgb is Hgb A 95-98% of adult Hgb is Hgb A  Remaining amount is A Remaining amount is A2 2 and A and A3 3 3
• 4.
  Sickle Cell Hemoglobin(Hgb S) Sickle Cell Hemoglobin (Hgb S)  The amino acid glutamic acid is replaced by The amino acid glutamic acid is replaced by amino acid valine amino acid valine  Ability of RBCs oxygen carrying capability Ability of RBCs oxygen carrying capability is impaired is impaired  Two possible genotype Two possible genotype – AS (sickle cell trait) AS (sickle cell trait) – SS (sickle cell disease) SS (sickle cell disease) 4
• 5.
  Pathophysiology of SickleCell Pathophysiology of Sickle Cell  Sickle Cell trait Sickle Cell trait – Clinical manifestation Clinical manifestation  Sickle Cell disease Sickle Cell disease – Clinical manifestation Clinical manifestation 5
• 6.
  Peripheral Blood Picture PeripheralBlood Picture  Varying number of sickled forms Varying number of sickled forms  Marked anisocytosis, poikilocytosis, Marked anisocytosis, poikilocytosis, and polychromasia and polychromasia  Target cells, nucleated RBCs, Target cells, nucleated RBCs, increased WBCs and platelets increased WBCs and platelets  Reticulocyte count is elevated: Reticulocyte count is elevated: (5-25%) (5-25%) 6
• 7.
  Sodium metabisulfite Sodium metabisulfite Principle Principle – Cells containing Hgb S will “sickle” Cells containing Hgb S will “sickle” when placed under a condition of when placed under a condition of reduced oxygen tension reduced oxygen tension Sodium Metabisulfate 15 minutes & 30 minutes 7
• 8.
  Dithionite tube test Dithionitetube test  Principle Principle – Hgb S is insoluble in a phosphate buffer Hgb S is insoluble in a phosphate buffer »Results in turbidity Results in turbidity – Hgb A is soluble Hgb A is soluble »Results in clear appearance Results in clear appearance – Other abnormal hemoglobins Other abnormal hemoglobins may result in turbidity may result in turbidity 8
• 9.
  ESR Clinical Significance ESRClinical Significance • The ESR is a The ESR is a nonspecific nonspecific test that test that suggests the possibility of an inflammatory suggests the possibility of an inflammatory process or tissue damage in the body process or tissue damage in the body • The ESR is The ESR is not diagnostic not diagnostic but is useful in but is useful in following the course of some diseases like following the course of some diseases like rheumatic fever, arthritis, and Lupus rheumatic fever, arthritis, and Lupus
• 10.
  Principle Principle  Based onprinciple of Based on principle of sedimentation where solids settle sedimentation where solids settle to the bottom of a liquid to the bottom of a liquid  A known quantity of A known quantity of anticoagulated blood is left anticoagulated blood is left undisturbed, the RBCs separate undisturbed, the RBCs separate from the plasma and fall to the from the plasma and fall to the bottom of the container bottom of the container
• 11.
  Procedure Procedure  Anticoagulated bloodis placed in a calibrated tube Anticoagulated blood is placed in a calibrated tube of standard dimensions of standard dimensions  It is incubated in a vertical position and left It is incubated in a vertical position and left undisturbed for a designated period of time undisturbed for a designated period of time (usually 1 hr) (usually 1 hr)  Distance RBCs fall within the given time is Distance RBCs fall within the given time is measured in millimeters and reported measured in millimeters and reported mm/hr mm/hr
• 12.
  Types of ESRMethods Types of ESR Methods  Wintrobe – simplest and Wintrobe – simplest and mostcost effective mostcost effective  Westergren – more sensitive Westergren – more sensitive  Automated methods Automated methods
• 13.
  Factors affecting ESR Factorsaffecting ESR  Plasma factors Plasma factors  Abnormal plasma proteins Abnormal plasma proteins may lead to rouleaux may lead to rouleaux formation formation (falsely increases ESR) (falsely increases ESR)  The single most important The single most important factor in determining factor in determining ESR rate ESR rate
• 14.
  RBC Factors RBC Factors Size Size – Large erythrocytes settle faster than Large erythrocytes settle faster than smaller ones and will increase ESR smaller ones and will increase ESR  Shape Shape – spherocytes settle rapidly while sickle cells spherocytes settle rapidly while sickle cells settle very slowly settle very slowly  Number of RBCs Number of RBCs – In anemia there are less RBCs and more In anemia there are less RBCs and more plasma volume which increases ESR plasma volume which increases ESR
• 15.
  Technical Factors Technical Factors Tilting Tilting  Noise and vibration Noise and vibration  Temperature Temperature  Test set up within 2hrs of blood collection Test set up within 2hrs of blood collection  ESR tube ESR tube  Well mixed sample Well mixed sample  Air bubbles Air bubbles  Accurately timed Accurately timed