Reticulocyte Count: Name: - SGD GRP: - Tests On Blood Cell Disorders

Name:_______________________________ SGD Grp:______
TESTS ON BLOOD CELL DISORDERS
RETICULOCYTE COUNT
I. Clinical Significance:
As a red cell matures, several days are needed for the hemoglobin containing cell to rid
itself of residual cytoplasmic ribonucleic acid after its nucleus has been expelled. The RNA
containing cell is slightly larger than the mature cell; it contains miscellaneous fragments of
mitochondria and other organelles as well as ribosomal RNA. These young red blood cells,
called “reticulocytes”, can often be distinguished on Wright stained peripheral smears by
their larger size and slightly gray or purple or bluish-pink appearance (polychromasia) due
to residual RNA.
The reticulocyte count is an index of the production of mature red cells by the blood
forming organs, mostly the bone marrow. Increased reticulocyte counts mean an
increased number of RBC is being put out into the peripheral blood in response to a
stimulus. Reticulocytes normally make up 1% of the total RBC count, but may exceed
levels of 4% when compensating for anemia. In exceptionally great reticulocyte responses
(reticulocytosis), there may even be nucleated RBC pushed out into the peripheral blood
due to massive red cell production activity of bone marrow.
* Immature precursor of the red blood cell, in which the remains of the residual RNA are
visible as network of filaments and granules (reticulum).
• very few are seen in normal blood as they are retained in the marrow until mature
• On remission of anaemia, when there is a high rate of production, reticulocytes appear
in the bloodstream (reticulocytosis).
• =On increased erythropoiesis, younger reticulocytes, also called "shift cells" are
released, which require two days to mature.
*Absolute Reticulocyte count

= The absolute reticulocyte count can also distinguish between hypo/hyperproliferative
anemia.
= If the absolute reticulocyte count is 100,000 /mm3 or higher, the anemia is
hyperproliferative type (i.e. hemolytic anemia or anemia of acute blood loss).
= If it is less than 100,000/ mm3 the anemia is hypoproliferative (iron, B12, or folic
deficiency, anemia of chronic disorder etc.).
II. Procedure:
1. Mount the prepared Blood Smears (stained with New Methylene Blue or Cresyl blue)
2. For counting, choose an area of the film where the cells are undistorted, mostly
one-cell layer, and where the staining is good. The cells should not overlap.
3. Cells are counted using the oil immersion lens.
4. Count a total number of 1000 red cells. As this number is being counted, jot down
the number of cells with granules or filaments seen.
 Report as percent (%) Reticulocytes counted (per 1,000 Red cells).

 Normal adult range: 0.5-1.5%
III. Calculation:
_____Total number of reticulated cells X 100 = % reticulocytes

1,000
Absolute count:
RBC/cumm X percent reticulocyte
 Absolute reticulocyte count:

Actual number of reticulocytes (range 25,000 – 125,000/ul)
IV. Questions:
1. Draw a Reticulocyte.
2. What is the reticulocyte percentage of your blood sample? Show your calculation.
3. What other red cell cytoplasmic inclusion bodies may be confused with reticulocytes.
How could they be differentiated from reticulocytes?
4. If a markedly anemic patient has a very low count or even absence of reticulocytes,
what is it indicative of?
Name:____________________________________ SGD No. ________ Date:_____________
X-CHROMOSOME (BARR BODY) DETERMINATION

TEST ON GENETIC DISORDERS
I. Clinical Significance:
“Barr bodies” are the morphological expression of the inactivated X chromosome.
“Drumstick”, as it is also called because of its shape, is an expression of an X-chromosome in
cells in which more than one are present, thus it is seen in normal women. “Drumsticks” in
Neutrophils and “Barr bodies” in epithelial cells are equivalent structures.
One Barr body can be seen in up to 3% of the neutrophils on a female's peripheral blood
slide. Recognition of a Barr body in a neutrophil is important in order to avoid reporting the
Neutrophil as hypersegmented.
Barr body (named after discoverer Murray Barr), the inactive X chromosome in a
female somatic cell, is rendered inactive in a process called lyonization.This has also been used
to investigate a sex-linked genetic disorder, such as multiple X chromosomes in a cell.
* The inactive ‘sex chromatin’ comprises of two superficially dissimilar structures:

(1) the Barr body, present in epithelial and other tissue cells;
(2) the drumstick of the polymorphonuclear leucocytes.
* Drumsticks are attached to the nuclear lobe by a single narrow stalk:

= which distinguishes them from other thicker projections, sometimes referred to as
"clubs."
* “Clubs” have a thicker, and sometimes, a double-stalk. This projection can be seen in
both males and females and has no clinical significance.
* ‘Drumsticks’, must also be distinguished from hair-like projections sometimes seen in the
band form, following irradiation or in patients with a malignant tumor that has
metastasized.
* Barr bodies in epithelial cells are present as a condensed mass against the inner nuclear
membrane.
*In rare chromosome disorders in which three or more X chromosomes are

present:
= two to three ‘Drumsticks’ per neutrophil can be seen.
* For example, patients with Klinefelter's syndrome (47,XXY) have an incidence of
drumsticks, although it is lower than that found in normal females.
* Clinical Significance:
1) Identify the gender of patients with ambiguous genitalia.
2) Investigate a sex-linked genetic disorder.
3) Barr body testing as confirmation in athletic competitions. (Olympics)
4) Medical and criminal forensics - in determining and legally defining the sex of an
individual.
II. Instruments & Materials:
Prepared Blood Smears
Immersion Oil
Microscope
Drumstick (Barr Body) on a Neutrophil
Barr Body in an Epithelial Cell Nucleus
III. Procedure:
1.Using the 100x oil objective (hpf), place a small drop of oil on the slide and
examine the smear for 100 Neutrophils.
11. Note all the Drumsticks identified in the 100 Neutrophils and record as
percentage.
> Normal Results

One Barr body in 1%- 3% of the neutrophils on a normal female's peripheral blood slide.
IV. Questions:
1) Record the percentage of Drumsticks seen: _________
2) Draw one of the Neutrophils you’ve seen with its ‘Drumstick’.
3) State whether the following Genetic Disorder would present Barr bodies or Not. Specify
the gender as well as the quantity of Barr bodies respectively.
Gender Presence of Barr Quantity of Barr

Bodies bodies/cell
a) Turner’s Syndrome = ______ ______________ ___________

b) Klinefelter’s Syndrome =_____ ______________ ___________
c) Mosaic individual = ______ ______________ ___________
(with XX & XY cells)

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Name:_________________________ SGD Grp:

TESTS ON BLOOD CELL DISORDERS

*Absolute Reticulocyte count

 Report as percent (%) Reticulocytes counted (per 1,000 Red cells).

_____Total number of reticulated cells X 100 = % reticulocytes

 Absolute reticulocyte count:

X-CHROMOSOME (BARR BODY) DETERMINATION

* The inactive ‘sex chromatin’ comprises of two superficially dissimilar structures:

* Drumsticks are attached to the nuclear lobe by a single narrow stalk:

*In rare chromosome disorders in which three or more X chromosomes are

Drumstick (Barr Body) on a Neutrophil

Barr Body in an Epithelial Cell Nucleus

> Normal Results

Gender Presence of Barr Quantity of Barr

a) Turner’s Syndrome = ___

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