Bone Marrow Examination Procedures Quiz

Bone Marrow Smear Preparation, Staining, and Examination

• Bone marrow examination is a crucial diagnostic tool in hematology.
• Bone marrow examination involves the analysis of bone marrow samples obtained through aspiration and biopsy.
• The aspirate yields a semi-liquid bone marrow sample, suitable for light microscopy, flow cytometry, chromosome analysis, or PCR.
• A trephine biopsy provides a solid bone marrow sample for microscopic examination, including immunohistochemistry, for cellularity and infiltrative processes.
• Bone marrow procedures are ordered based on clinical symptoms and peripheral blood findings.
• Bone marrow puncture is contraindicated in patients with severe bleeding disorders (e.g., hemophilia, vitamin K deficiency).
• Common complications are mild soreness lasting 12-24 hours after the procedure.

Objectives of Bone Marrow Examination

• At the end of a bone marrow examination, a pathologist can:
• Discuss the clinical conditions requiring bone marrow examination.
• Identify suitable bone marrow aspiration sites for various age groups.
• Describe the techniques for preparing and examining bone marrow smears.
• Evaluate bone marrow cellularity.
• Determine the myeloid-to-erythroid ratio.
• Perform a differential count on bone marrow smears.

Bone Marrow Aspiration vs. Biopsy

• Both procedures are invasive and require technical skill, but aim for different outcomes.
• Aspiration collects bone marrow fluid, used for identifying cell types and morphology. It's typically simple, safe, and less painful than a biopsy.
• Biopsy collects a solid bone marrow sample that allows assessment of cellularity and architecture, and can detect focal lesions, granulomas, multiple myelomas and other diseases.

Bone Marrow Specimen Collection

• Typically done through the posterior iliac crest and less frequently on the sternum or other sites.
• Different needles (e.g., Salah, Klema, Jamshid) are used based on the nature of the procedure and the patient.
• Procedures are performed under sterile conditions, often in an operating theatre.

Indications for Bone Marrow Examination

• Unexplained anemia and cytopenia/pancytopenia.
• Mulitlineage abnormalities of circulating blasts in adults.
• Abnormal peripheral blood smear morphology.
• Diagnosing, staging, and monitoring hematologic malignancies.
• Investigating suspected bone marrow metastases.
• Unexplained focal bony lesions or unexplained organomegaly/mass lesions.
• Microbiological evaluations for fever of unknown origin or infections.
• Evaluation of iron stores.
• Monitoring after chemotherapy, radiation therapy, or a stem cell transplantation.

Bone Marrow Films

• Preparing bone marrow films involves spreading the sample on glass slides and staining with Romanowsky dyes.
• Longer fixation times (e.g., at least 20 minutes in methanol) are necessary for high-quality bone marrow staining.
• Using Perl's method can help assess iron content.

Particle/Crush Smears

• Particle/crush smears involve placing bone marrow particles between slides and pulling them apart.
• This method provides an accurate reflection of bone marrow morphology.
• Squashing and smearing can cause cellular disruption and distortion, impacting the accuracy of the study.

Systemic Scheme for Examining Aspirated BM films

• Low power (x10): Assess cellularity and megakaryocytes, identify clumps of abnormal cells, and assess macrophages.
• Higher power (x40-x100): Identify stages of hematopoiesis in myeloid and erythroid lineages, calculate M:E ratio, perform differential counts, observe for areas of necrosis, and evaluate iron content.

Cellularity of Bone Marrow

• Cellularity is assessed relative to the patient's age and site of aspiration and compared to areas occupied by fat spaces and nucleated cells.
• Normal bone marrows are noted as either normocellular or normoplastic.
• Higher values than expected for a patient indicate hyperplasia, which can be seen in conditions like polycythemia.
• Lower values indicate hypoplasia and are associated with marrow failure (e.g., aplastic anemia, myelofibrosis).

Myeloid-to-Erythroid Ratio (M:E)

• This ratio describes the relationship between myeloid and erythroid cells in bone marrow.
• In normal adult bone marrow, myeloid cells outnumber erythroid cells.
• An increase or decrease in this ratio can indicate a specific disease or condition.

Differential Count on Aspirated Bone Marrow

• The assessment of cell types as percentages is not routinely performed, due to the time involved and its limited clinical value.
• Irregular cell distribution and inclusion of peripheral blood cells can affect the reliability of the count.
• A minimum of 200 cells should be studied if performing differential counting.

Terminal Deoxynucleotidyl Transferase (TdT)

• TdT is a nuclear enzyme marker for primitive lymphoid cells, used to study lymphoblastic leukemia.
• High levels are common in lymphoblastic leukemias, less common in acute myelogenous leukemias (AMLs).

Molecular Methods (FISH)

• Newer DNA probes and PCR techniques are utilized to rapidly confirm chromosomal abnormalities and monitor residual diseases.
• FISH (fluorescence in situ hybridization) identifies specific chromosome translocations associated with particular leukemias.

Systemic Lupus Erythematosus (SLE)

• SLE is a chronic rheumatic autoimmune disease.
• The immune system attacks its own tissues, leading to inflammation, widespread inflammation and tissue damage.
• Commonly it is seen in childbearing-age women and may include clinical features like skin rash, arthralgia, fever, renal, cardiac and vascular lesions.

LE Cell Phenomenon

• LE cells are abnormal cells produced in SLE.
• They are characterized by an immunoglobulin of the IgG, IgM or IgA class that causes the depolymerization and phagocytosis of the nuclear chromatin by an intact polymorph.
• The distinguishing features of the LE cell in the cellular morphology include:
  • Ingested nucleus or cellular materials.
  • The bulk of the cell is occupied by a spherical, homogeneous mass that stains purplish brown.
  • The polymorph's lobes are wrapped around the ingested material.
• LE cell detection requires preparing specific types of bone marrow slides and requires expertise.

Demonstration of LE Cells

• Identifying LE cells involves examining films, especially their edges and tails, for at least 10 minutes and analyzing (minimum of 500 polymorphs).
• Dead nuclei in films may heighten suspicion but aren't diagnostic on their own.
• Differentiating LE cells from "tart cells" (monocytes with ingested material) is essential for proper diagnosis

Method of LE Cell Detection (Rotary Method)

• A method for detecting LE cells using a patient's blood sample.
• The procedure involves using heparinized blood, glass beads, and a centrifugation step to prepare a smear from the blood.
• The smear is then prepared for staining and examination.

"Tart Cells"

• Tart cells are monocytes or histiocytes with engulfed viable nuclei.
• These are not equivalent to LE cells, and don't provide any diagnostic value.
• Critical for distinguishing from LE cells, which can be confused with "tart cells."