An immature red blood cell without a nucleus, having a granular or reticulated appearance when suitably stained.
Reticulocytes are the immature RBC that contain nucleus.
They are originally seen at the site of their formation i.e. bone marrow. They take 2-3 (lays for maturation only about 1-2% of circulating RBCs are Reticulocytes.
3. Introduction:
Definition: An immature red blood cell without a
nucleus, having a granular or reticulated appearance
when suitably stained.
Reticulocytes are the immature RBC that contain
nucleus.
They are originally seen at the site of their formation
i.e. bone marrow. They take 2-3 (lays for maturation
only about 1-2% of circulating RBCs are Reticulocytes.
4. Aim: To determination of Reticulocytes count.
Clinical Significance: The number of Reticulocytes in peripheral
blood is a reflection of red cell forming activity of bone marrow.
Increase in Reticulocytes count indicates increased activity of the
bone marrow. This is known as Reticulocytosis. It is seen in
Haemolytic anaemia or acute blood loss. Low count of
Reticulocytes indicates suppression of bone marrow. It is seen in
aplastic anaemia.
Method: Supra vital method
5. Normal Range:
Adults: - 0.2 to 2 %
Infants: - 2 to 6 %
Principle: Supravital staining method is used for
Reticulocytes count. Blood is mixed with the stain & the
stain enters the cells in living condition. The RNA in the
cells is precipitated by staining, as dark blue network.
Blood smear is made afterwards. Since direct count is
not possible, a relative count is taken against the
6. Requirements:
1. Glass slides
2. Test tube
3. Pasteur pipette
4. Microscope
5. Staining solution :(Staining solution contain
Brilliant cresyl blue (1.0g), Na citrate (0.4g), NaCl
(0.85g) & distilled water (100ml)
Dissolve first Na citrate in Na chloride & then the
dye. Filter it & store in a plastic container.
Specimen: - EDTA blood sample or capillary
heparinized blood. Perform the test within 2 to 3 hr
of blood collection.
7. Procedure:
1.Identify the right patient.
2. Collect all the requirements.
3. Do the labelling.
4.Collect required amount of blood by vein or capillary puncture.
5. Filter a small amount of the stain.
6. In a labelled test tube take two drops a blood & two drops stain by
using two separate Pasteur pipette.
7. Mix properly then cover the tubes with cotton plug & keep it at 37
degrees for 30 minutes.
8. Prepare a thin smear of the stained blood specimen.
9. Air-dry the smear.
10. First examine the smear under the low power than change it to
the oil immersion objective.
11. Reticulocytes are identified by line, deep violet filaments &
granules arranged in a network. Red cells stain pale blue.
8. Calculation:
Example: Total no. of red cell counted = 1500
No. of reticulocytes seen = 75
Reticulocytes % = 75 x 100
--------------
1500
= 5%
9. Additional Information:
1. Staining time should not be less than 30 min.
2. Mix the blood & the stain gently but thoroughly prior to
making the smear. This is imp. The Reticulocytes have a
lower specific gravity than maturated cells & therefore settle
on top of the red cells in the mixture. Thus an unmixed or
poorly mixed blood specimen may not give the true picture.
3.Red cells showing highly refractive areas may be
confused with Reticulocytes. These artefacts in the red cells
are probably due to moisture in the air & poor drying of the
