BONE MARROW PUNCTURE ( BMP ) Prof.

Adi Koesoema Aman SpPK(KH)

Divisi Hematologi Departement Patologi KLinik FK USU / RSUP.H.A.Malik , Medan

BONE MARROW PUNCTURE ( BMP ) . 1. Aspirasi . - Red Maroow . 2. Aspirasi & Biapsi . - Structure Tulang . - Trabeculae Tulang . - Sinusoid . - Jaringan hematopoitik dan non-hematopoitik

1 . Aspirasi : - Jarum Salah .

2. Aspirasi Biopsi : - Jarum Silvermman . - Islamic Needle .

Bone Marrow -Definition

Bone refers to location of bone marrow after birth Marrow: Latin for middle. Hence Bone Marrow is located in middle of bone.

DISTRIBUTION OF BONE MARROW

Pelvis-40% Vertebrae-28% Cranium-mandible-13% Ribs-8% Sternum-2% Ends of long bones-8%

TYPES OF MARROW
Yellow marrow (inactive) composed primarily of fat

Red marrow (active in hematopoiesis)

Marrow puncture needle - Salah

X- sternal aspiration sites

Tibial aspiration site

Posterior iliac crest aspiration

BM SITES AND INDICATIONS FOR ASPIRATIONS Posterior iliac crest (any age, for aspiration and biopsy) Anterior iliac crest (only for infants and children, for aspiration and/or biopsy) Sternum (unsuitable under 15 y/o;only for aspiration) Tibia (infants below 18 mo.; only for aspiration)

Scheme for examining BM aspirate films

1) Determination of cellularity 2) Identification of megakaryocytesNote morphology and maturation sequence 3) Look for clumps of abnormal cells 4) Identify macrophages Examine for evidence of haemophagocytosis , malaria

pigment, bacterial or fungal inf (x 40 , x 100)

5) Identify all stages of maturation of myeloid and erythroid cells 6) Determine myeloid:erythroid ratio 7) Perform a differential count 8) Look for areas of BM necrosis 9) Assess the Iron content of macrophages

Normal cellularity

Hypocellular

Hypercellular

Staining of cells
Identification of various types of blood cells

CYTOLOGICAL EXAMINATION OF BONEMARROW

Myeloblast

Promyelocyte

Neutrophilic myelocyte

Metamyelocyte

Band granulocyte

segmented granulocyte

Band granulocyte

segmented granulocyte

Leukemia cells

LYMPHOCYTE

SERIES

Lymphoblast

Prolymphocyte

Lymphocyte

Lymphocytic leukemia cells

MONOCYTE SERIES

Monoblast

Promonocyte

Monocyte

PLASMA SERIES

proplasmacyte

plasmacyte

Plasmablast

MEGAKARYOCYTE SERIES

Megakaryoblast

promegakaryocyte

Granular megakaryocyte Thrombocytogenous megarkaryocyte

Other cells

Organization of Alkaliphilic cells

Fibroblasts

Mesh phagocytes

Fat cells

Endothelial cells

Osteoblasts

Osteoclasts

PENGENALAN SEL HEMATOPOITIK

UNTUK MENGENALI JENIS SEL DARAH DAPAT DILAKUKAN DGN CARA :
1. MORFOLOGI SEL . 1.1. PENGECATAN RUTIN . 1.2. PENGECATAN SITO-KIMIAWI . 2. PENENTUAN CLUSTER DIFFERENTIATION ( CD ) FLOWCYTOMETRI IMMUNO PHENOTYPING

1.1. PENGECATAN RUTIN .

PENGECATAN RUTIN DILAKUKAN DG CARA : 1. MGG ( MAYGRUNWALD GIEMSA STAIN . 2. ROMANOWSKY STAIN .

1.2. PENGECATAN SITO-KIMIAWI

PENGECATAN SITO - KIMIA DIGUNAKAN UNTUK DIAGNOSE BANDING DAN KLASIFIKASI LEUKEMIA , YAITU UNTUK : 1. MEMBEDAKAN MIELOBLAST DAN MONOBLAST . 2. MENENTUKAN SEL LIMFOBLAST . 3. MEMBEDAKAN LEKOSITOSIS DGN REAKSI LEKEMOID . 4. MEMPELAJARI KELAINAN ATAU DEFISIENSI ENZYM DLM NEUTROFIL

Cyto - Staining methods

POX-NAENAPPAS Fe dyes

(Peroxidase, POX)
Principle:
Benzidine + H2O2
[O]

Aniline blue + Nitroso sodium cyanide

Granules (azurophilic) are stained

Cells of Myeloid series - POX Positive

Cells of lymphocytic, megakaryocytic , erythroid series &

Monocytes - POX Negative

POX Reaction

Leukemia cells Leukemia cells

Lymphocytic leukemia cells

Sudan black B
Demonstrates lipoproteins and lipids Principle : Sudan black B (Fat soluble dye)
Intracellular protein bound lipids

- Black intracytoplasmic granules

- Diffuse black staining of cytoplasm

Positive reaction

Sudan black B positive

- Pro , neutrophilic and meta myelocytes - Band cells - Segmented neutrophils Sudan black B negative

- Lymphocytes
- Megakaryocytes - Erythroblasts - Normoblasts - Platelets

Neutrophil alkaline phosphatase (NAP)

Naphthol phosphate
NAP

Naphthol
Diazonium salt

Bright Red color

Insoluble precipitate

NAP

present in mature neutrophils

Other blood cells negative

Chronic myeloid leukemia (NAP)

Acid phosphatase (ACP)

Principle : Specimen + Naphthol phosphate substrate

Naphthol

Diazonium salt
Colored insoluble precipitate (red granules) (at the sites of acid phosphatase activity)

Cells of hairy cell leukemia and auer bodies Strongly positive Granulocytes - Positive RBCs - Negative

Hairy cell leukemia

Auer bodies

Naphthol AS-D chloroacetate esterase (AS-D NCE)

AS-D naphthol insoluble red ppt.

Naphthol AS-D chloroacetate

AS-D NCE H2O

+ Corinth v salt

Mast cells , granulocytes and their precursors (band & juvenile cells an promyelocytes)

Positive

 -Naphthol acetate esterase (- NAE)

Principle similar to AS-D NCE - Naphthyl acetate is used as a substrate and Pararosaniline as coupling azo dye Enzymatic activity results in intracytoplasmic dark red granules

Periodic acid schiff stain (PAS) Cytochemical stain (PAS) demonstrates Glycogen
Glycogen Periodic acid Oxidation Parafuchsin(schiff reagent) Aldehydes

Red color compound

Stains demonstrating Iron (Gomori prussian blue stain)

Common blood disorders

Anemia Leukemia Myelodysplastic syndrome Multiple myeloma Primary ITP

MA KASIH IYA . SAYA DI SURAMADU SURABAYA