Bone marrow aspiration & trephine biopsy, Complication of BM Aspiration, Clinical significance, Indication of Bone Marrow Aspiration and Biopsy, Types Of Needles, Site for Bone Marrow Biopsy And Aspiration, types Of Smear for Bone Marrow, Advantages of Bone Marrow Trephine Biopsy

1. PRESENTED BY:-
Sanjeev Kumar
BSC MLT 1
DEPARTMENT OF HAEMATOLOGY

2. INTRODUCTION
BONE MARROW ASPIRATION
 Aspiration is aspirate of the contents of bone
marrow drawn under pressure by puncturing
marrow cavity.
 When there is abnormal finding in routine
blood film, bone marrow evaluation become
important to know the exact cause.
TREPHINE BIOPSY
 Removal of a small core of BM under
local anesthetic. It is used to assess
BM structure, the no. & distribution of
all types of blood cell .
 Trephine biopsy is 1-2cm long 2-3mm
in dia. Of the marrow containing bone
taken by rotating specialized needle
under pressure
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3. BONE MARROW ASPIRATION
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4. Historical Events
1868- Neuman and Bizzazero sequeezed
the marrow from rib of human cadaver.
1929- Arinkins proved marrow aspiration
as a safe easy and useful technique.
1876- Mosler use a regular wood drill to
obtain marrow particle.
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5. Indication for B M aspiration
 Determine the cytological types and proportions of
haematopoietic cells in the marrow
 Study and classify nucleated blood cells of the bone marrow
 Determine the ratio of immature white blood cells to red
blood cells (M:E ratio, myeloid:erythroid precursors)
 Assess number and morphology of megakaryocytes
 Investigate anaemia (abnormally low number of red blood
cells in the bloodstream)
 Evaluate abnormalities in the blood’s ability to store iron
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6.  Diagnose an infection with an unknown cause, blood
disorders, chromosomal or genetic diseases, cancer of the
blood cells, such as leukaemia, lymphoma, multiple
myeloma, neutropenia (low neutrophils count),
thrombocytopenia (low platelet count), Polycythaemia (high
red blood cell count), pancytopenia (reduction in the number
of red and white blood cells, as well as platelets)
A bone marrow aspiration may be performed in people
with certain types of cancer to:
monitor the side effects of chemotherapy
see whether the treatment is working.
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7. requirements
 Aspiration needle
 Should be stout and made of hard stainless steel.
 About 7-8cm in length with a well fitting stylet provided with a
adjustable guard to prevent over penetration.
 The most commonly used needle is Salah / Klima.
 Bore size 0.2mm.
 Spirited swab
 Betadine
 Local anaesthesia (2% lignocain and Xylocaine)
 Two 10-20ml syringe
 Clean grease free and iron free glass slide
 Smooth edge glass slide spreader to make aspirate film.
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8. Types of Needles
a) Salah Needle: has a guard with
a side screw
b) Klima Needle: has a guard
which screw along the length of
the needle. The needle is
advanced into the marrow
cavity with the stylet locked is
place, the stylet is then removed
and syringe used to aspirate the
marrow.
c) Jamshidi Needle: which is
longer can be used for both
aspiration & biopsy from iliac
crest.
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Salah Needle
Klima Needle

9. Anticoagulant for different studies
• Place at least 0.5ml of the
bone marrow aspirate in
Sodium Heparin tube and
EDTA tube.
Chromosome/PCR/
Cytogenetic studies
• Place as much bone marrow
aspirate as possible into a
sterile EDTA tube and mix.
For Flow
Cytometry
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10. fixative
For slides
For Aspirated
Histology
Section
• Absolute
Methanol
• Absolute
ethanol +15%
formalin (1:1)
1) For Slides
ABSOLUTE
METHONOL
2) For Aspirated
Histology Section
Absolute ethanol
+15% formalin (1:1)
FIXATIVE
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11. Sites for BMA
1. Iliac spines or crest:This is the most commonly employed site
for reason of safety, has a large reservoir of marrow, a decreased risk of pain and
accessibility. There is no large blood vessels or nerve to this area.
2. Sternum:Last resort in those older than 12 year. Avoided since causes more
pain for the patient and present the risk of heart injury.
3. Tibia:Sampled only for infants younger than one year.
4. Spinous process of lumber vertebra:This is an
additional site for aspiration in adults.
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12. Site for Bone Marrow Aspiration
 Posterior Superior iliac crest:- This
is the most commonly employed site
for reason of safety, a decreased risk of
pain and accessbilty.
 Anterior Superior iliac Crest:- This
is an alternate site when the posterior
ilia crest is unapproachable or not
available due to infection, injury or
morbid obesity.
Aspirate only
 Sternum:- Last resort in those older
than 12 year. Avoided since causes
more pain for the patient and present
the risk of heart injury.
 Tibia:- Sampled only for infants
younger than one year.
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13. 13

14. PROCEDURE
 The patient is placed in the lateral position, with the top leg flexed and
the lower leg straight.
 Palpate the iliac crest, and mark the preferred sampling site with a
pen.
 Aseptic technique is employed, including sterile gloves and gown.
 The site is prepared with an antiseptic (eg, povidone-iodine or
chlorhexidine gluconate), scrubbed, and draped, exposing only the site
to be sampled.
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From Posterior iliac crest (PIC)
Skin preparation

15. • The skin and the underlying tissue to the periosteum are
infiltrated with a local anesthetic (eg, approximately 10
mL of 1% Xylocaine).
• A 10-mL syringe with a 25-gauge needle is used to inject
an initial 0.5 mL directly under the skin, raising a wheal.
• A 22-gauge needle is used to penetrate deeper into the
subcutaneous tissue and the underlying periosteum, an
area roughly 1 cm in diameter.
Local anaesthetic injection
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16. •Adequacy of the anesthesia is tested by gently prodding the
periosteum with the tip of the needle and questioning the patient for
any painful sensation.
•Once the needle contacts the bone, it is advanced by slowly rotating
clockwise and anticlockwise until the cortical bone is penetrated and
the marrow cavity is entered. Contact with the marrow cavity is
usually noted by a sudden reduction in pressure. The depth of the
penetration should not extend beyond an initial 1 cm.
Aspiration needle placement
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17. •Once within the marrow cavity, the stylet is removed.
Using a 20 mL syringe, approximately 0.3 mL of bone
marrow is aspirated. The material collected for bone
marrow slides is generally not mixed with an anticoagulant,
and it is processed immediately; this avoids any cellular
morphologic artefacts.
Bone
marrow
aspiration
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18. 18
Riskinvolved in bonemarrowaspiration
Risk Involved In Bone Marrow Aspiration
 People with bleeding problems have a higher chance of bleeding
from biopsy site. If patients have bleeding problems, pressure
will be put on the biopsy site for at least 10 minutes after the
biopsy. In rare cases, patients may be given a blood product
(clotting factor or platelets) in a vein in arm before the
aspiration to prevent bleeding after the aspiration/biopsy.
 Infection of skin or the bone at the biopsy site, especially in
people with compromised immune system.
 Injury to heart, a lung, or a major blood vessel if the sample is
taken from the sternum. This complication is very rare.
 Pain after the procedure.

19. Examination of Aspirated Bone Marrow
1) Direct smears
3) Particle
smear
4) Imprint smear
2) Concentration smear
5) Histological
section
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20. DIRECT
Clean grease free and
Iron free slide
Slide put in slant
position then put the
aspirate, wait few
seconds to drain out of
blood from the
aspirate or suck off
with the help of
Pasteur pipette or
filter paper or put in
watch glass then take
particle and make
smear.
Air dry and fix in
methanol.
PARTICLE
Pick up one or more
particle from
aspirate put on the
slide and cover with
another slide press
gently and pull
slight apart
longitudinally
IMPRINTS
One or two slides are
gently touched in
several places by the
exposed marrow at the
surface of the biopsy.
These imprints are
allowed to dry, the
slides are then fixed &
stained in the same
manner as film.
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21. Preparation of aspirated
smear
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22. CONCENTRATION
1. Put the anticoagulated marrow
aspirated material into plastic tube
centrifuge at 1500 rpm for 20-
25min.
2. Take Buffy coat layer mix with
equal volume of plasma.
3. Then make smear. 22

23. STAINING
ROUTINE STAINS SPECIAL STAINS
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Romanowsky Stain’s
May Grunwald Giemsa stain
Giemsa stain
Leishman stain
Haematoxylin & Eosin stain
Perl’s stain
Reticulin stain
Cytochemical stain (for
Leukemia classification)

24. Advantages of Marrow Aspiration
It provides a semi
quantitative
assessment of the
blood cell precursors.
It is an easy, safe and
inexpensive means to
find out the
haemopoietic
abnormalities
It provides the
diagnosis of several
hereditary and
acquired bening
malignant disease
chromosome study.
It provides response of
the treatment of
luekemia and some
lymphoma.
Useful to assess the state
of iron-stores and
metabolic disorder that
affect macrophages such
as Gaucher disease.
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25. Disadvantages of Marrow Aspiration
The arrangement of cells with in the marrow and the relationship of
one cell to another more or less destroyed by the process of aspiration.
In fibrotic marrows, little blood is also aspirated.
Dry tap i.e. when there is little or no marrow aspirated.
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26. TREPHINE BIOPSY
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27. Indications of BM Biopsy
 Repeated failure to obtain adequate material by aspiration.
 In myelofibrosis biopsy for confirmation as fibrosis cannot be
demonstrated with certainty on smears from aspirates.
 In case of metastatic tumor, lymphoma or granulomatous disease
may be observed in aspirated material, but they are easily found in
sections from biopsy specimens.
 An end point of aplasia.
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28. REQUIREMENTS
1. Disinfectant: 70% alcohol.
2. Local anthesia: 2% Xylocaine
3. Fixatives: 95% Zenker sol, 10% Formal saline,
10% NBF
4. Gloves
5. Clean slides
6. Biopsy needle:
 Jamshidi needle
 Turkel & Bethell
 Vim-Silverman needle
 Islam trephine
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29. Jamshidi Trephine
It consist of a cylindrical needle of 2mm bore size constant
except tapered concentrically at the distal position. Ending in a
sharp beveled cutting tip with a precisely fitting stylet which
extends 1-2mm beyond the length of needle.
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30. ISLAM TREPHINE VIM-SILVERMAN
 It has a core securing device which
obtain a uniform core of marrow
without disturbing morphology &
anatomy
 It consists of an outer cannula, an
obturator, and an inner split needle with
longitudinal grooves in which the tissue is
retained when the needle and cannula are
withdrawn.
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31. SITES FOR BM TREPHINE BIOPSY
 1. Anterior superior iliac spine.
 2. Posterior superior iliac
spine
(commonly used this site)
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Place for the procedure:-
Under aseptic
conditions(minor OT) .

32. PROCEDURE
Needle inserted in skin through the subcutaneous
tissue through cortex of the bone, which is sensed
by a decreased resistant to the forward movement
of the needle
Infiltrate the skin, subcutaneous tissue and
periosteum overlaying the selected site with local
anesthesia.
Disinfect the area with the disinfectant. (large
area of skin at the site of procedure)
Ask the patient to lie down in prone position or
side wise (left or right lateral).
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33. conti…….
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The obturator is then removed and the
needle is slowly advanced with
reciprocal, clockwise-counterclockwise
twisting motions around the long axis.
After sufficient penetration of bone , the
needle is rotated several times in its axis
and withdraw about (2-3mm).
The needle is then reinserted to
the original depth at a slightly
different angle and rotate
several times in order to free the
specimen from attachment in
the marrow cavity.

34. Determining length of biopsy core in needle by
carefully reinserting obturator.
Ideally, the core length should be 2 cm or greater.
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35. The needle is
slowly withdrawn
using the same
twisting motion
employed during
insertion.
The core of marrow
inside the needle is
removed by inserting the
probe through the
cutting tip and extruding
the specimen through
the hub of the needle.
Make touch
preparation before
fixing the specimen
in 95% Zenker’s
fluid or 10% formal
saline.
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36. POST BIOPSY CARE
 FINISHING UP:-
 After procurement of the marrow specimens,
bleeding must be stopped, the procedure site
must be cleaned up, needles properly
disposed of in a Sharps container, and the site
bandaged.
 A procedure note must be placed on the
patient’s chart.
 After procedure is completed
apply pressure over the posterior
Ilium for about 60 min.
 Pressure is obtained by a
pressure dressing & by having
the patient lie recumbent(flat
position) in bed.
 After that assuming no bleeding
is observed the patient can getup
& go about their normal
activities .
 Patients are advised to avoid
washing the procedure site for at
least 24 hr after the procedure is
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37. TREPHINE PREPARATIONS
Touch smear:-
 Collected specimen extruded in rough the hub of needle &
touched gently across the slide by using the forceps.
 Put the smears in the fixative jar.
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 Touch smears have special
value because cells of primitive
type tend to be present in
compact clusters and thus may
be missed if only the more fluid
portions of the marrow are
examined.

38. Histology section preparation:-
 Fixation
 Decalcification
 Processing
 Sectioning
 Staining
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39. ADVANTAGES OF TREPHINE
BIOPSY
It gives cell
and stroma
constituent.
In Cases of
dry tap
Represent
all cells
types.
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40. Precautions of BONE MARROW
 Bone marrow procedure is preffered with strict attention to
ASEPTIC technique & UNIVERSAL precautions for the
transmission of BLOODBORNE pathogens.
 Fixatives fluids, slides, tubes with the proper
ANTICOAGULANT should be at bed side so the specimen
can be preserved immediately & slides preparations that are
of suitable quality.
 ALLERGIES or previous adverse reaction to medicine
should be discussed with doctor.
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41. Conti…
 Any med. Including Herbal or Nutritional
supplements should be evaluated to interfere with
proper coagulation(clot formation).
 Caution should be used when the Herbs gingto,
garlic, have been utilize as supplement due to risk
of bleeding.
 Pregnancy, lactation & pre-exesting, platelets or
blood disorder should be evaluated before procedure
is undertaken.
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