Manual of basic techniques for a health laboratory - libdoc.who.int

9. Haematology 2719.3 Estimation of the haemoglobin concentrationHaemoglobin is the red pigment contained in erythrocytes. It consists of proteinchains and iron-containing molecules.Units of measurementThe SI unit for expressing haemoglobin concentrations is millimole per litre (mmol/l). When this unit is used, it is necessary to specify the chemical structure to whichit applies. In practice, this means that the term “haemoglobin(Fe)”should be used instead of the simple term “haemoglobin”. However,as an interim measure, before making the change to millimoleper litre, some laboratories are using the unit “gram per litre” (g/l).When this unit is used, the simple term “haemoglobin” suffices, andit is not necessary to say “haemoglobin(Fe)”. Values in grams perlitre may be converted into values in millimoles per litre by multiplyingby 0.062.Example:haemoglobin 150g/l ¥ 0.062 = haemoglobin(Fe) 9.3mmol/lIn this manual calculations and values are usually expressed in bothforms. It should be noted that if the unit “gram per litre” is used, thevalues are 10 times greater than values in the traditional unit “gramper 100ml”. For example, 150g/l = 15.0g/100ml.Fig. 9.21 Transferring the blood to a specimentube9.3.1 Haemiglobincyanide photometric methodPrincipleThe blood is diluted in Drabkin diluting fluid, which haemolyses the red cells andconverts the haemoglobin into haemiglobincyanide (cyanmethaemoglobin). Thesolution obtained is examined in a spectrophotometer (or colorimeter). Its absorbanceis proportionate to the amount of haemoglobin in the blood.The haemiglobincyanide photometric method gives the most accurate haemoglobinestimations. It should be used wherever possible.Materials and reagents● Spectrophotometer 1 (or colorimeter)● Spectrophotometer (or colorimeter) cuvettes● Test-tubes● Test-tube rack● Blood (Sahli) pipettes, 0.2ml● Drabkin diluting fluid (reagent no. 21)● Reference solution, which may be:— the fresh haemiglobincyanide reference solution used to calibrate theinstrument,— a reference solution previously calibrated against the haemiglobincyanidereference solution, or— a blood sample of known haemoglobin concentration.A calibration curve must be prepared before the spectrophotometer (or colorimeter)can be used for haemoglobin estimation. From such a curve a graph can beprepared and a table made for the haemoglobin values.1Some spectrophotometers run either on mains electricity or on current from a motor car battery.One model is supplied by UNICEF: reference no. 09.309.98 (110V battery) or 09.310.00(220V battery); it can be ordered from the following address: UNICEF, UNICEF Plads, Freeport,DK 2100 Copenhagen, Denmark.