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Hemoglobin concentration

Hemoglobin concentration. Hematology Lab. Hemoglobin concentration. Hgb concentration estimated by: light intensity, or color matching technique * Spectrophotometer * Methods used are: Hemoglobin cyanide method : Reference method

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Hemoglobin concentration

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  1. Hemoglobin concentration Hematology Lab

  2. Hemoglobin concentration Hgb concentration estimated by: light intensity, or color matching technique *Spectrophotometer* Methods used are: Hemoglobin cyanide method : • Reference method • Internationally recommended for Hgb concentration measurement Oxyhemoglobin method

  3. The absorbance is measured using spectrophotometer of 540 nm wave length. • Then it is the concentration determined using standard curve.

  4. Principle of Hgb cyanide method • Blood is diluted in a solution: • contains potassium cyanide + potassium ferricyanide (Drabkin solution) • As a result this reaction will occure: • Hgb, HbCO  methemoglobin HiCN (cyanmethemoglobin) • HbS will not be converted using this method

  5. Methods 1:201 dilution (blood: diluents) • 20µL of blood in 4 mL Cover and invert the tube several times • Let it stand at R.T for 5-10 minutes to insure complete conversion • Pour into cuvette and read absorbance at 540nm wave length spectrophotometer. • Set spectrophotometer to 100% transmittance at 540 wave length, using cyanmethemoglobin as blank. • Continue reading patient samples and record % transmittance or absorbance. • Determin Hgb value of sample using, Beers law and STD curve.

  6. Results and Calculations • HiCN is used as standerd • To calculate the Hgb concentration use the following equation: • A of test sample x concentration of STD x dilution factor A of Standard 1000

  7. Standard curve A Test Absorbance Hgbconcentration glL Testconcentration

  8. Linearity of standard curve • To ensure linearity of standard curve: • 3 STD are used • 1:1 dilution 20µL blood : 20µL diluent • 1:2 dilution 20µL blood : 40µL diluent • 1:3 dilution 20µL blood : 60 µL diluent

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