Examination of Bone Marrow

• Bone marrow is the site of hematopoiesis in postnatal life.

• Bone marrow is one of the largest organs in the body, and although
distributed widely in the skeleton.
• It is located within the cavities of the bones and mainly consists of
hematopoietic cells, vascular sinusoids, fibroblasts, fat cells, and
macrophages.
• There are no lymphatic channels in the bone marrow
• There are two types of marrow: red and yellow.
• Red marrow refers to the active hematopoietic tissue while fatty
tissue comprises the yellow (inactive) marrow
• Red coloration is due to presence of large amounts of developing red
cells.
• The average volume of bone marrow (red and yellow) in an adult is
3000-4000 ml.
• Red or active marrow constitutes 1500 ml.
Structure of Bone Marrow

Bone Marrow consist of

• Vessels
• Nerves
• Hemopoitic cells
• Reticuloendothelial cells
• Fat tissue
• stroma
• After development, mature blood cells leave the bone marrow and
enter the circulation by passing through or in between the
endothelial cells of the sinusoids.
There are two techniques for sampling of bone marrow: aspiration and
biopsy.
• In aspiration, bone marrow fluid is obtained by a special needle and
syringe. Smears from this material are prepared on glass slides,
stained, and examined under the microscope.
• In bone marrow trephine biopsy (core biopsy), a small tissue piece of
bone marrow is removed with a special needle, processed to obtain
histological sections, and examined.
INDICATIONS FOR BONE MARROW EXAMINATION

• Before performing bone marrow aspiration/biopsy, one should assess

clinical features, treatment received, and relevant laboratory test
results (especially basic blood studies and peripheral blood smear).
• Based on above data, if appropriate indication exists, then
examination of bone marrow is carried out and findings are
correlated to arrive at the final diagnosis.
INDICATIONS FOR BONE MARROW ASPIRATION
1.Unexplained cytopenia:
If the cause of cytopenia (anemia, leukopenia, or thrombocytopenia) is
not apparent from blood investigations and clinical details, bone
marrow examination is indicated.
(A) To distinguish amongst causes of microcytic hypochromic anemia
(e.g. iron deficiency from chronic disease), bone marrow
examination can be done to assess storage iron.
(B) In leukopenia or thrombocytopenia, bone marrow examination is
helpful in distinguishing peripheral destruction from deficient
production.
2. Suspected acute leukemia:
In majority of cases, acute leukemia can be diagnosed from
examination of a blood smear and cell counts. Bone marrow
examination is carried out in acute leukemia for:
• Detection of subleukemic or aleukemic leukemia
• Comparison of baseline marrow smear with follow-up marrow
aspiration smears during treatment
• Diagnosis of acute myeloid leukemia with trilineage dysplasia
• Cytogenetic analysis.
3.Suspected myelodysplastic syndrome.
4. Suspected myeloproliferative disorders including chronic myeloid
leukemia, polycythemia vera, essential thrombocythemia, and
myelofibrosis.
5. Suspected plasma cell dyscrasia: Bone marrow aspiration is indicated
if multiple myeloma is suspected from clinical and radiologic features.
6. Suspected chronic lymphoid leukemias like chronic lymphocytic
leukemia, prolymphocytic leukemia, hairy cell leukemia, etc.
7. Investigation of pyrexia of unknown origin:
Sometimes, bone marrow aspiration smears are helpful in detecting
Histoplasma capsulatum and Leishmania donovani organisms in
macrophages of bone marrow. Aspirated material can also be cultured
to isolate above organisms or mycobacteria.
8. Suspected storage disorder like Gaucher’s disease or Neimann-Pick
disease.
9. Suspected infections like kala azar, miliary tuberculosis, or
histoplasmosis.
INDICATIONS FOR BONE MARROW BIOPSY

1. Repeated failure of aspiration (dry tap) which may be due to faulty

technique, myelofibrosis or leukemia.
2. Suspected aplastic anemia
3. Suspected myelofibrosis.
4. Suspected focal lesions like granuloma, metastatic deposit, or
infiltrate of lymphoma.
5. Suspected hairy cell leukemia.
6. Suspected bone disorder, e.g. osteopetrosis.
7. Staging of lymphoma.
CONTRAINDICATIONS
• Bone marrow aspiration or biopsy is contraindicated in hemophilia
and other coagulation disorders; however in these patients it can be
performed under cover of appropriate replacement therapy.
• Thrombocytopenic purpura is not a contraindication since applying
firm pressure for 5 minutes at the puncture site can prevent excess
bleeding.
Sites
Iliac spines or crest:
The most frequently used site for both marrow aspiration and biopsy
in children (>1 year of age) as well as in adults is posterior iliac crest
(posterior superior iliac spine).
This site has a large reservoir of marrow, is located just beneath the
skin and therefore easily accessible. Also, there are no large blood
vessels or nerves close to this area, and as the patient’s back is towards
the physician, patient’s apprehension is less.
In obese patients, anterior superior iliac spine, being more easily
localized, can be used
• Sternum:
Previously, sternum was commonly used for aspiration of bone marrow
in adults (at the level of second intercostal space in midline). However,
it is associated with the risk of perforation of posterior sternal plate
and puncturing of underlying large blood vessels and right atrium with
serious consequences.
It also causes greatest patient anxiety. This site should not be used in
children as the bone is thin and marrow cavity is small.
• Spinous processes of lumbar vertebra:
This is an additional site for aspiration in adults.
• Tibia:
In infants under 1 year of age, marrow can be aspirated from the
medial aspect of upper end of tibia just beneath tibial tuberosity.
Bone Marrow Aspiration Method

• An informed consent should be obtained before the procedure.

• Bone marrow aspiration or biopsy should be performed by the
physician. An assistant is required for preparation of smears.
• A sterile tray should be prepared containing autoclaved bone marrow
aspiration needle, sterile disposable syringes with needles, local
anesthetic solution, clean and dry glass slides, spreader slide, gloves,
drapes, gauze, and a skin antiseptic solution. All aseptic precautions
should be observed during the procedure.
Bone Marrow Tray
• For aspiration from posterior superior iliac spine, patient should lie on
one side with back towards the physician, knees and hips flexed, and
the knees drawn towards the chest. The site for aspiration should be
selected and scrubbed with soap and water.
• After wearing sterile gloves, antiseptic solution is applied in a circular
fashion, moving from center towards periphery.
• A sterile drape is placed over the area with its central opening over
the aspiration site.
Patient position (superior posterior iliac
crest).
Site preparation
• Skin and periosteum are infiltrated with a local anesthetic.
• First inject beneath the skin surface and advancing the needle
further, a larger amount is injected into the periosteal surface.
Local anesthetic injection
• After waiting for 5 minutes for anesthesia to take effect, bone
marrow aspiration needle is inserted along with the fitted
stylet. (Stylet prevents blockage of lumen of needle by tissues
through which needle passes).
• When the bone is reached, the needle is rotated clockwise and
anticlockwise and slowly advanced into the bone, maintaining
steady and firm pressure.
• When the marrow is reached, a slight “give” (decrease in
resistance) will be noted. . If the needle is placed correctly, it
will be fixed by the surrounding bone and will remain rigid and
unmoving.
Aspiration needle placement
• A 5 or 10 ml syringe is attached to the needle and a small
amount of marrow is aspirated (till the first drop of blood
appears i.e. 0.25-0.50 ml) by quickly pulling the plunger of the
syringe.
• Aspiration of larger amount of blood causes dilution of
marrow sample by peripheral blood with subsequent
difficulties in interpretation of smears. If no material is
aspirated, stylet is replaced, needle is redirected, and
aspiration attempted again.
Bone marrow aspiration
• The syringe should be handed over to the assistant for
preparation of smears on glass slides.
• The smears should be made promptly, before clotting occurs,
by putting one drop of the aspirated material near one end of
a glass slide and spreading it similar to a blood film.
• If immuno- phenotyping or cytogenetic analysis is to be
carried out, further marrow sample should be aspirated in a
second syringe and dispensed in a tube containing heparin
anticoagulant.
Bone marrow aspiration and biopsy slide
preparation
Bone marrow aspiration and biopsy slides before
staining
• After completion of aspiration, the stylet should be reinserted
into the needle and the needle is removed. Sterile gauze is
placed over the site and light pressure is applied till bleeding
ceases. A larger dressing is then applied.
 Bone Marrow Biopsy Method
• Preparation of the patient and local anaesthesia are similar to
aspiration.
• Percutaneous trephine biopsy of bone marrow is commonly
obtained from posterior superior iliac spine.
• Jamshidi or Islam needles are commonly used.
• Often, bone marrow aspiration and biopsy are combined
together; aspiration is carried out first followed by biopsy.
• If both aspiration and biopsy are combined, then, after aspiration,
either (i) the needle is advanced a little further (1-3 cm) into the
bone, or (ii) the needle is withdrawn and reinserted through the
same skin incision but placed at a different site in the bone (about
1 cm away).
• For biopsy, the needle should be advanced through the bone
rotating it clockwise 10 times. The needle should be removed by
anticlockwise rotation. The biopsy should be removed gently from
the hub end of the needle by inserting the stylet through the point
of the needle.
• For adequate assessment, biopsy should measure at least 1.6 cm in
length. Biopsy should be placed in a fixative solution (either 10%
formalin or preferably Helly’s fluid).
• Dressing should be applied to the site similar to aspiration.
Bone marrow biopsy - needle placement
Bone marrow biopsy specimen
Bone marrow biopsy specimen in fixative solution
• Along with bone marrow
aspiration/biopsy, peripheral
blood smears should be
prepared from finger prick and
venous blood should be
collected in EDTA anticoagulant
for cell counts.
 COMPLICATIONS OF BONE MARROW ASPIRATION
AND/OR BIOPSY
1. Local infection: This complication, which is more likely to occur
in neutropenic patients, can be prevented if strict aseptic
precautions are observed.
2. Haemorrhage: Serious haemorrhage can occur if (i) marrow
biopsy is done without adequate replacement cover in
coagulation disorders, and (ii) great vessels or heart is injured
during sternal aspiration.
3. Cardiac tamponade or mediastinitis: This is likely if posterior
sternal plate is penetrated during sternal aspiration.
References
• Kawthalkar SM. Essentials of Clinical Pathology. New
Delhi: Jaypee Brothers Medical Publishers (P) Ltd, 2010.
Thank You