Hemoglobin and hematocrit determination
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Hemoglobin and hematocrit determination
- 1. Colorimetric Determination of Hemoglobin Indicator of Fluidity and Transport Capacity Baldeo, Biendima, Go, Olivar, Soriano
- 2. L-R: Pipette Sahli’s Standard Hemometer Tube Stirring Rod Dropper
- 3. Hemoglobin Determination 0.1 M HCl at 2% mark 20 mm3 blood Hemolysis of RBC releasing hemoglobin which is then converted to acid hematin (Ferriheme chloride)
- 4. Add drops of water until colors matched
- 5. Results Normal Levels: Men: 14-17 g/100 mL Women: 12.5-15 g/100mL Infants: 17-23 g/100 mL Volunteer Hemoglobin Content (g/100 mL) Alyssa 13.2 Pat 13 Cyndi 22 Kass 14.2
- 6. Conditions Involved • Anemia • Hypoxia • Leukemia • Cirrhosis • Hypothyroidism • Myeloma
- 7. Limitations to Sahli’s Method • Standard is not permanent • There is considerable delay in the development of the permanent color (many instances reach 20 mark) – 10 minutes: 95% acid hematin – 20 minutes: 98% acid hematin – 1 hour: 100% acid hematin • Large errors have been found in Sahli’s pipette, recalibration is needed before using them
- 8. Improving Sahli’s Method • Cyanomethemoglobin absorbance reading at 540 nm • Blood samples treated with: – Potassium ferricyanide – Potassium cyanide – Potassium dihydrogen phosphate
- 9. Total Oxygen-Carrying Capacity (TOC)
- 10. TOC Volunteer Hemoglobin Content (g/100 mL) TOC (mL O2) Alyssa 13.2 563.36 Pat 13 473.51 Cyndi 22 996.424 Kass 14.2 556.57 Normal levels: Male: 20 mL O2/ 100 mL blood Female: 19 mL O2/ 100 mL blood
- 11. Hematocrit Determination Indicator of Blood Viscosity
- 12. Centrifuge blood samples in Eppendorf tubes for 10 min
- 13. Hematocrit Determination H1 H2 Where: • H1 = height of the RBC column • H2 = height of the RBC + height of the plasma column • Calculate Hc% (hematocrit) value
- 14. Results Volunteer Hematocrit (%) Alyssa 106.67 Pat 54 Cyndi 57 Kass 33 Normal Values: Children, 9-11 years (whole blood): 34%-43% (0.34-0.43) Females, 12-14 years (whole blood): 34%-44% (0.34-0.44) Males, 12-14 years (whole blood): 35%-45% (0.35-0.45) Females, 15-17 years (whole blood): 34%-44% (0.34-0.44) Males, 15-17 years (whole blood): 37%-48% (0.37-0.48)
- 15. Hematocrit • Normal hematocrit values are: – Newborn: up to 60% – Adults: (males): 40- 54% – (Females): 36 – 46% • Pregnancy: decreased hematocrit, especially in the last trimester as plasma volume increases • Children: varies with age • Hematocrit = 3x hemoglobin for normal RBC count and hemoglobin amount only
- 16. Hematocrit
- 17. Hematocrit and Blood Viscosity
- 18. Conditions
- 19. Conditions • Elevated hematocrit – Sign of dengue shock syndrome. – Polycythemia vera – bone marrow produces excessive numbers of RBCs – Chronic obstructive pulmonary diseases – Hypoxia – Capillary leak syndrome – Dehydration (hypovolemia)
- 20. Conditions • Decreased Hematocrit – Hemorrhage – Chronic kidney disease – Inadequate iron uptake – Iron deficiency due to blood loss during menstruation – Anemia – Malnutrition – Overhydration
- 21. Conditions • A hematocrit of less than 15% can result in cardiac failure • A hematocrit of over 60% may result in spontaneous blood clotting
- 22. Technical Errors • Results increased in : – Strenuous exercise – High altitude • Technical errors – Inadequately mixed blood sample – Length of time spent and speed of centrifuge – Prolonged venous status during venipuncture
- 23. Normal RBC Count = Normal Hematocrit = Normal Hemoglobin Content? • The NORMAL HEMOGLOBIN level does NOT ensure NORMAL RBC COUNT and HEMATOCRIT. • NORMAL HEMATOCRIT does NOT ensure NORMAL HEMOGLOBIN content or NORMAL RBC COUNT.
- 24. Normal RBC Count = Normal Hematocrit = Normal Hemoglobin Content? • Not a definitive test – For example, if you're otherwise healthy and have no signs or symptoms of illness, results slightly outside the normal range on a complete blood count may not be a cause for concern, and follow- up may not be needed.
- 25. Blood Indices Combining Results for RBC count, Hemoglobin Determination, and Hematocrit Determination
- 26. Mean Corpuscular Volume (MCV)
- 27. Results Volunteer Hematocrit (%) RBC count (millions/mm3) MCV (femtoliters) Alyssa 106.67 6,360,000 176.67 Pat 54 8,330,000 64.83 Cyndi 57 4,500,000 126.67 Kass 33 4,315,000 76.48 Normal range: 90-95 µm3 = 90-95 femtoliters
- 28. HIGH MCV • Liver disease due to alcohol abuse • Myelofibrosis • Reticulocytosis • little vitamin B12 or folic acid • Megaloblastic Anemia
- 29. LOW MCV • Anemia
- 30. Mean Corpuscular Hemoglobin (MCH)
- 31. Results Volunteer Hemoglobin (g/100mL blood) RBC count (millions/mm3) MCH (picograms) Alyssa 13.2 6,360,000 20.75 Pat 13 8,330,000 15.61 Cyndi 22 4,500,000 48.89 Kass 14.2 4,315,000 32.91 Normal range: 27-33 picograms
- 32. Mean Corpuscular Hemoglobin Concentration (MCHC%)
- 33. Results Volunteer Hemoglobin (g/100mL blood) Hematocrit (%) MCHC (g/100mL) Alyssa 13.2 106.67 12.37 Pat 13 54 24.07 Cyndi 22 57 38.60 Kass 14.2 33 43.03 Normal range: 30-34 g/100mL
Notas del editor
- Hb is a chromoprotein, hence the Hb-content in a blood sample may be determined by measurement of its colour.
- conversion of hemoglobin by hydrochloric acid into acid hematin, which has a brown color in solution
- The intensity of the color is related to the amount of hemoglobin in the blood sample.The more hemoglobin, the more water required to obtain a color match.
- Anemia – physiological incapability to replace old RBCs; fatigue, uncontrolled bleeding, higher infection rate; low O2 content in blood to tissues = hypoxiaLeukemia = cancer of lymphatic system and blood-producing tissues (marrow); excessive and abnormal WBCs produced, crowds out RBCs and platelets; low proprotion of hemoglobin.Cirrhosis – liver scarring by hepatitis or alcohol abuse, irreparable; Bile is a yellow-green pigment produced from breakdown of hemoglobin; no bile production = excessive hemoglobin leftover in liver.Hypothyroidism = Insufficient hormone produced by thyroid; eythropoeitin in bone marrow produces RBCs stimulated by thyroid hormones.Myeloma – Kahler’s disease; excessive plasma levels = high antibody production, resulting in low hemoglobin readings in blood <10 g/dL
- GQ2Acid hematin is not stable and the color gradually starts fading.10 mins minimum
- GQ 5
- volume of erythrocytes expressed as a percentage of volume of whole blood in the sample. Also known as packed cell volumeHematocrit separates blood in a tube by centrifugation into: 55% Plasma a straw colored fluid Serum different from plasmaFormed elements [buffy coat, (<1% leucocytes and platelets)] Formed elements [45% erythrocytes]
- The viscosity of blood increases drastically as the hematocrit increasesThe viscosity of whole blood at normal hematocrit is about 3 -three times as much pressure is required to force whole blood as to force water through the sameblood vessel. When the hematocrit rises to 60 or 70, which it often does in polycythemia, the blood viscosity can become as great as 10 times that of water, and its flow through blood vessels is greatly retarded.
- Polycythemia – retarded flow of blood in BV; High hematocrit
- MCV can be falsely elevated in the presence of red blood cell agglutination (as incold agglutinin disease or paraproteinemia) or severe hyperglycemia (glucose > 600mg/dL) as red blood cells become swollenMacrocytic – large RBCS; Microcytic – small; Normocytic - normal
- Liver Disease – deposition fats / phospolipids in circulating RBCsScarring of the bone marrow means the marrow is not able to make enough blood cells. As a result, the liver and spleen try to make some of these blood cells. This causes these organs to swell ineffective in removing wastes from RBCsOver production of reticulocytes which are 20% larger than normal RBCs (patients with haemolytic anemia)Needed for DNA synthesisImpaired DNA synthesis unaffected RNA synthesis; slow division but rapid deposition of cellular components
- normal amount of hemoglobin (normal MCHC) are called normochromic.When the MCHC is abnormally low they are called hypochromic, and when the MCHC is abnormally high, hyperchromic.