Date: _________ Lab 06 Student Id:

__________

OBJECT- Estimation of Hemoglobin percentage in human blood by Sahli’s method

Apparatus/chemicals
N/10 hydrochloric acid
Distilled water
Hemoglobin meter
Lancet
Introduction
Hemoglobin, also spelled hemoglobin is an iron-containing protein in the blood of many
animals- in the red blood cells (erythrocytes) of vertebrates that transports oxygen to the tissues.
Hemoglobin forms an unstable, reversible bond with oxygen; in the oxygenated state it is called
oxyhemoglobin and is bright red; in the reduced state it is purplish-blue. It is made up of four
polypeptide chains (a1, a2, B1, and ẞ2). Each chain is attached to a heme group composed of
porphyrin (an organic ring-like compound) attached to an iron atom. These iron-porphyrin
complexes coordinate oxygen molecules reversibly, an ability directly related to the role of
hemoglobin in oxygen transport in the blood.

Hemoglobin develops in cells in the bone marrow that become red blood cells. When red cells
die, hemoglobin is broken up: iron is salvaged, transported to the bone marrow by proteins called
transferrin’s, and used again in the production of new red blood cells; the remainder of the
hemoglobin forms the basis of bilirubin, a chemical that is excreted into the bile and gives the
feces their characteristic yellow-brown color.
Function
Hemoglobin is a protein that is used to carry oxygen through the bloodstream from the lungs to
the tissues. This is important for survival. Hemoglobin has a lower affinity for oxygen the lower
the concentration of oxygen gets. This has great implications for the human body and has helped
us adapt very effectively. The lower affinity and lower concentrations mean that when we are
working out, our bodies are low on oxygen which means hemoglobin has less affinity for oxygen
and can more easily drop the oxygen off at human tissues.
Normal hemoglobin levels
Gender
Male-------Level------14-18g/dL.
Females-------12-14g/dL.
Indications of Hemoglobin Estimation
 To determine the presence and severity of anemia
 Screening for polycythemia
 To assess response to specific therapy in anemia.
 Estimation of red cell indices
 Selection of blood donors.
Sahli's Method:
Sahli's hemoglobin meter is a manual device that contains a hemoglobin tube, pipette, and stirrer,
as well as a comparator. Hydrochloric acid converts hemoglobin to acid hematin, which is then
diluted until the color of the solution matches that of the comparator block. The Sahli
haemometer method utilizes the conversion of hemoglobin 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. Water is added to dilute the brown solution until it matches that of a standard. The more
hemoglobin, the more water is required to obtain a color match. Hemoglobin values are read at
the meniscus of the brown solution.

Procedure:
 The Sahli's Method was used in the lab to measure hemoglobin levels.
 Fill Sahli's Hb tube up to the 2g mark with HCl solution.
 Prick the fingertip and draw 20µL of blood directly from the fingertip into the pipette.
 Add the blood to the Hb tube with the HCl in it.
 Stir with a stirrer until the solution's color is uniform.
 Add distilled water a few drops at a time until the color of the solution matches the color
on the comparator.
  Once it matches, note the reading on the Hb tube; that is the hemoglobin level in g/dL.

Observation:
___________________

Discussion:
______________________________________________________________________________
______________________________________________________________________________
______________________________________________________________________________
______________________________________________________________________________
______________________________________________________________________________
______________________________________________________________________________
______________________________________________________________________________
______________________________________________________________________________
______________________________________________________________________________
______________________________________________________________________________
______________________________________________________________________________
______________________________________________________________________________
______________________________________________________________________________
______________________________________________________________________________
______________________________________________________________________________
______________________________________________________________________________