Estimation of haemoglobin concentration of blood - Practical

Estimation of haemoglobin concentration of blood

Haemoglobin is the main component of the red blood cells & helps in the transport of O₂ and CO₂. Haemoglobin is a conjugated protein that consists of a basic protein, the globin and haeme. Each hemoglobin molecule consists of one molecule of globin and four molecules of haeme each containing a molecule of iron.

Aim

To measure the haemoglobin content of your blood using hemo meter.

Apparatus and reagents required

Sahli-Hellige Hemometer, N/10 HCl, distilled water, lancet, cotton swabs, alcohol, dropper.

Description of apparatus

The Sahli-Hellige Hemo meter has the following components:

Comparator : It accomodates a glass comparison tube which is graduated in gram percent of Hb (2-24) as well as percentage (20-140). This tube is used for holding the acid hematin. The comparator is provided with non-fading standard brown tinted glass plates.

Haemoglobin pipette: A pipette showing a single mark, indicating 20 cu mm. volume is provided.

Glass stirrer : Used for stirring the contents of the comparison tube.

Procedure

Add N/10 HC1 to the comparison tube upto the 20% mark.

Prick the ball of a finger using a sterile lancet after wiping with cotton dipped in alcohol, to obtain blood.

Wipe off first one or two drops of blood.

When a good-sized blood drop forms, suck the blood into the haemoglobin pipette upto the 20 cu.mm mark.

Wipe away the excess of blood adhering to the tip of the pipette with cotton.

Immediately transfer the contents of the pipette into the tube containing HCl solution.

Suck a small amount of HCl acid into the pipette and blow it into the tube.

Mix and allow the tube to stand for 10 minutes.

Add a few drops of distilled water and stir the contents with a glass-stirrer.

Continue to add water drop-by-drop and stir the contents each time until the colour of the solution is just darker than the colour of the standard one. Note the reading, against diffused light.

Again, add one or two drops of distilled water & when the colour matches exactly with the standard , note the reading, against diffused light.

Again, add one or two drops of distilled water- so that, the colour of the solution at this time becomes lighter than the standard solution. Note this reading, against diffused light.

Take the average of the three readings.

Result: The haemoglobin content is _______________ gms/100ml of blood.

Precautions

Wipe off the blood adhering to the tip of the pipette, since such a blood can give a false high value of Hb. The blood drop obtained by squeezing the finger or from a cold and cyanosed finger is likely to give high values of Hb. Hence this should be avoided.

The colour of the standard tinted glass should be checked periodically against the standard acid hematin solution.

The acid-blood mixture must be kept for atleast 10min before the dilution, to cause a complete conversion of Hb into acid hematin. The normal range of haemoglobin is as follows,

Men-13.5 - 18.0 gm/100ml.

Women - 11.5 - 16.5 gm/l00ml

Infants - 13.6 - 19.6 gm/100 ml.