Chapter 1 Rodak Overview of Laboratory Hematology PDF

Hematology MTEC 406 Maria de los A. Oliveras, MHSA, MT(ASCP) Medical Technologist Professor UAGM-Gurabo Campus  Hematology CELL COUNTING CELL MORPHOLOGY CLOTTING FACTORS  Chapter 1 Overview of Clinical Laboratory Hematology...

Hematology MTEC 406 Maria de los A. Oliveras, MHSA, MT(ASCP) Medical Technologist Professor UAGM-Gurabo Campus  Hematology CELL COUNTING CELL MORPHOLOGY CLOTTING FACTORS  Chapter 1 Overview of Clinical Laboratory Hematology ◦ Define Hematology, Hematopoiesis, Erythropoiesis, Objectives Myelopoiesis ◦ Describe two main differences between band and segmented ◦ It will distinguish between different neutrophils compounds in the blood, including ◦ Distinguish between eosinophils and basophils platelets, white and red blood cells. ◦ Describe four morphological traits of monocytes. ◦ It will detail the recommended methods of analysis for the ◦ List four characteristics of lymphocytes. evaluation of each hematopoietic ◦ Compare a large lymphocyte to a monocyte. component, their expected and ◦ Explain the process of releasing platelets from the bone clinical values. marrow.  Hematology Blood Study and Its Related Disorders  Hematology Blood: ◦ Transports Oxygen – Lungs to Tissue ◦ Cleans fabrics by removing CO2 ◦ Transports glucose, proteins, lipids ◦ Moves waste to the liver and kidneys ◦ Plasma, among many components, provides the clotting enzymes that protect blood vessels from trauma and maintain circulation. ◦ Transports and nourishes Blood Cells: ◦ Three categories of cells: Erythrocytes, Leukocytes, Thrombocytes  Hematology Blood: Hematology is the study of blood in Health and Disease. This includes problems with: Red Blood Cells, White Blood Cells, Platelets, Blood Vessels, Bone Marrow, Lymph Nodes, Spleen, Proteins involved in Bleeding and Clotting.  Hematology Blood: ◦ Hematology focuses primarily on the study of blood cells: Erythrocytes, Leukocytes, Thrombocytes ◦ Through the: ❖Staining, Counting, Analysis ❖Record of Morphology or Appearance, ❖Phenotype, Genotype, of these cells. Blood diseases and many Systemic Diseases can be predicted, detected, diagnosed and monitoring therapy. Blood: Hematology Hematology studies Hemostasis: A natural process that stops blood loss when damage occurs. This process involves: ❖ Vasoconstriction (vascular spasm) – endothelial or vascular tissue ❖ Plug formation (plug) of platelets ❖Coagulum Formation (Coagulation Proteins) Historia Hematology ◦ 1642- Anthon van Leeuwenhoek – noticed cells in the blood ◦ 1657- Athanasius Kircher- He described maggots in the blood ◦ 1674- Anthon van Leeuwenhoek – RBC Counting ◦ 1842- Albert Donne- Discover platelets ◦ 1846- George Gulliver- Difference between lymphocytes and granulocytes by size ◦ 1874- Louis Charles Malassez- WBC count by hemocytometry ◦ 1875- Georges Hayem- defines methods for counting platelets ◦ 1879- Paul Ehrlich- use aniline dye to dye WBC. ◦ 1902- James Homer Wright – developed the Wright Stain.  Hematology History ◦ 1953- First Automated Analyzer – Coulter Model A ◦ 1980- “Boom" Blood Cell Analyzers – Greater Accuracy and Precision ◦ 1982 - introduced the techniques of Molecular Biology ◦ 1985 – Molecular studies for hematological malignancies are introduced. ◦ 2010 - Molecular Biology Laboratory is established. ◦ 2012- new technologies are incorporated, cytogenetics in Advanced Technology Center ◦ 2014- la técnica FISH – “Fluorescence In Situ Hybridization” ◦ 2020- Real Time PCR: Molecular Technology Revolution  Hematology Blood Plasma Average blood volume: 4 – 6 L 91.5% Water ◦ Women: 4 – 5 L 8.5% Solutes ◦ Men: 5 – 6 L 55% albumin ◦ 8% of Total Body Weight 38% globulins ◦ Blood pH= 7.35 - 7.45 7% Fibrinogen ◦ 55% Plasma Other: electrolytes, hormones, non- ◦ 45% Cells or Elements protein nitrogenous compounds, ❖45% RBC nutrients, and respiratory gases. ❖< 1% WBC & Platelets  Blood Cells: Normal Values RBC WBC / Platelets ◦ Women ◦ WBC 4.2 – 5.4 x 1012 / L (SI) 4.8 – 10.8 X 109 / L 4.2 – 5.4 x 106/uL (convencional) 4.8 – 10.8 X 103 /uL ◦ Men ◦ PLTs 4.7 – 6.1 x 1012 / L 150 – 400 x 109 / L 4.7 – 6.1 x 106 /uL 150 – 400 x 103 / uL These reference values may vary by age, gender, geographic location, health status, or illness. CBC Hematology CBC Test “Complete Blood Cell” Performed by automated analyzers It includes specific measures to: ◦ Red Blood Cells, ◦ White Blood Cells and ◦ Platelets. CBC  Hematology CBC Test “Complete Blood Cell” Specimen – for Counting ◦Whole Blood for Counting ❖Capillary Blood Specimen - excellent for morphology: non-chemical alters cells. ❖Venous – Anticoagulant: EDTA ◦ Mixing well: Homogeneity ◦ 60 times reversal / 2 minutes automatic rotator ◦ Fresh Sample  Hematology CBC Test- Complete Blood Cell Specimen – for Counting EDTA - Ethylene Diaminetetraacetic Acid (K2EDTA) ◦ Minimal effect on cell morphology ◦ Process within 6 hrs. collection to RT – avoid false results ◦ Process within 24 hrs. collection at 4 C. ◦ Attention to the ratio of blood to anticoagulant Affects only HCT – falsely low values by shrinkage cells Heparin is unsatisfactory as an anticoagulant, especially leukocytes. Hematology: CBC Test The MT is responsible for the integrity of the specimen You must ensure that the sample: It has been collected in the correct tube/anticoagulant. It is clot-free It is hemolysis-free Sufficient volume to meet the criteria for anticoagulant ratio: sample. Be analyzed or prepared to be stored within the established time. Ensure an accurate analysis Complete Blood Count (CBC)  Automated Equipment ◦ Many blood cell analyzers also provide comments on RBC, WBC, and Platelet Morphology. ◦ When one of the results from the blood cell analyzer is abnormal, the instrument provides an indication of this, sometimes called a flag. ◦ In this case, a reflex Blood Film Examination is performed  CBC Composite of Cells Found in Peripheral Blood of Healthy Individuals. Erythrocyte (RBC) (A); Neutrophil (segmented neutrophil, polymorphonuclear neutrophil, PMN) (B); Band (band neutrophil (C); Eosinophil (EO) (D); Basophil (BASO) (E); Lymphocyte (Lymph) (F); Monocyte (MONO) (G); Platelet (PLT) (H). (Wright-Giemsa stain, ×1000.) RBC Red Blood Cell CBC Test RBC Counting (#) Anemia vs Polycythemia Automated Equipment ◦ Impedance Principle Electric Current ◦ Cells passing through an opening ◦ Where a stream is flowing. ◦ The cell causes an electrical resistance ◦ That is counted as voltage pulses. Electrical Impedance Coulter Principle Blood suspension diluted exactly with conductive and isotonic diluent to preserve the cell shape. A current flows through the opening between the E+ and E- electrodes Each cell passing through the opening increases electrical resistance and causes a voltage pulse. The pulse is proportional in height to the volume of the cell. CBC Test RBC Counting(#) Anemia vs Polycythemia Manual – It consists of three steps: 1. Sample Dilution-- Dilution with 0.85% saline --- 1:200 dilution ◦Normal saline is like blood osmolality ◦Retains intrinsic morphology: They do not swell or crenate 2. Measuring a Diluted Sample Volume 3. Count the cells in that volume-- Cells/Unit Blood Volume ◦ Expressed cubic millimeter (mm3) – linear dimension hemocytometer ◦ 1 mm3 = 1.000003 uL (conventional) Hemocytometer  Hemocytometer The areas for the standard white blood cell count are labeled W, The areas for the standard red blood cell count are labeled R. The entire center square, outlined in blue, is used for counting platelets. The side view of the hemacytometer shows a depth of 0.1 mm from the surface of the counting grid to the coverslip. mm3 CBC Test RBC Anemia vs Polycythemia Hemoglobin Main Component: Red Blood Cells Conjugated protein – O2 and CO2 transport vehicle Measurement Method: Cyanmethemoglobin (HiCN) Colorimetry /540 nm CBC Test RBC Anemia vs Polycythemia Hemoglobin- Cyanmethemoglobin (HiCN) ◦ Rx. Drabkin (potassium cyanide and potassium ferricyanide) ◦ Potassium ferricyanide oxidizes Hgb to Methemoglobin (Fe3+) (Hi) ◦ Potassium cyanide provides CN- ion to form HiCN----- maximum absorbance at 540 nm ◦ Standard Curve -- expresses grams/dl (g/dl) ◦ Source Error: Sample Turbidity -- Increases absorbance not by Hgb ◦ Hyperlipidemia ◦ High number of leukocytes (>30,000/mm3) ◦ Increase Abnormal Proteins in Plasma RBC Anemia vs Polycythemia CBC Test Hematocrit Erythrocyte Volume Ratio to Whole blood (Packed Cell Volume (PCV)) ◦ Reflects Red Cell Concentration (RBC) – not mass ◦ Expressed as a percentage (%). ◦ Anticoagulant: maybe EDTA o Heparin RBC Anemia vs Polycythemia CBC Test Hematocrit Methods: ◦ Direct Centrifugation: ◦ Macro methods (Wintrobe) ◦ Micro methods (Capillaries with heparin) ◦ Deliberate (Calculated) : Automated Equipment MCV x RBC  CBC Test RBC Anemia vs Polycythemia Hematocrit Methods: Direct Centrifugation: Direct Examination– Valuable Information -- Relative Height RBC, Buffy Coat, Plasma Buffy coat: grey-red layer between RBC and plasma (leukocytes, platelets) Plasma: orange or green – suggests increased bilirubin Pink or red plasma – suggests hemoglobinuria --- hemolysis Cloudy plasma – nephrosis, abnormal hyperglobulinemia – cryoglobulinemia. Source Error: ◦ Improper centrifugation: duration and/or speed ◦ Excess EDTA-low HCT due to crenate/RBC shrinkage; ◦ Not mixing the sample well, not reading HCT well RBC Anemia vs Polycythemia CBC Test Indices ---- Important for Classification Anemia ◦ MCV- Mean Corpuscular Volume - diameter formula: (HCT /RBC) x 10 ◦ femtolitres (fT) --- um3 (cubic micrometric ) = 10-15 L ◦ MCH- Mean Corpuscular Hemoglobin - Hemoglobin Content/Red Cell (rbc) ◦ formula:(HGB /RBC) x 10 Picograms (pg) = 10-12g ◦ MCHC- Mean Corpuscular Hemoglobin Concentration formula: (HGB / HCT ) x 100 ◦ Grams/dl (SI) / percent ( %) (Conventional Unit) ◦ Average hemoglobin concentration in each volume of “Packed RBC”. ◦ Reflects RBC stain intensity and RBC central pallor. ◦ RDW- Red Cell Distribution Width – Derived Histogram RBC ◦ CV RBC Distribution= (SD x 100)/ average/ Standard Deviation of RBC Volume. ◦ Degree of variability RBC volume– Anisocytosis RBC Prueba CBC Resume: Indices: ◦ MCV- Mean Corpuscular Volume - formula: (Hct /RBC) x 10 fL ◦ MCH- Mean Corpuscular Hemoglobin - formula:(Hgb /RBC) x 10 pg ◦ MCHC- Mean Corpuscular Hemoglobin Concentration formula: (Hgb / Hct ) x 100 % ◦ RDW- Red Cell Distribution Width – distribution MCV-- % CV(SD/average X 100) histogram RBC  WBC White Blood Cell CBC Test White Blood Cell (WBC) Count (#) ----- Infection "White Blood Cell" - Colorless cells in a suspension of unstained cells. Use acid solution to lyse rbc. RBC can hinder counting (500 -1000 x more) Automated Current Impedance Principle Manual- --- Hemocytometer -- diluted 1:20 Uses a lot to count cells Body fluids (CSF, Peritoneal, Synovial, Renal) CBC Test WBC Infection ◦ Differentiate Cells by Cell Type: ◦ Neutrophils/ Bands/ Eosinophils/ Basophils – Granulocytes ◦ Lymphocytes/ Monocytes – Mononucleates ◦ Absolute number / % of each fraction of the total WBC ◦ Leucopenia vs leukocytosis ◦ Leukemia – “Shift to the Left “  WBC Parameter Composite of Cells Found in Peripheral Blood of Healthy Individuals. WBC Neutrophil (segmented neutrophil, polymorphonuclear neutrophil, PMN) (B); Band (band neutrophil (C); Eosinophil (EO) (D); Basophil (BASO) (E); Lymphocyte (Lymph) (F); Monocyte (MONO) (G); (Wright-Giemsa stain, ×1000.) Platelets Plts CBC Test Platelets (Plts) Counting (#) ◦ Automated: Current impedance principle ◦ MPV- Average Platelet Volume Manual- Diluted Acid Solution 1:20 Function: ◦ Maintains the integrity of blood vessels and their repair - "Coagulation“ ◦ Clots – thrombosis ◦ Thrombocytopenia vs Thrombosis CBC Test Blood Smear Examination--- “Extendidos” ◦ Wright Stain – staining Romanowsky’s Method ◦ Basic dyes (Methylene Blue) and acids (Eosin) dissolved methyl alcohol solution ◦ Fixation and Staining ------ Light Microscope ◦ Estimate WBCs and Platelet counts. ◦ They analyze the Morphology RBC and WBC: ◦ Color, Size, Shape, Cytoplasmic inclusions, Condensation of the core. WBC Differential– Categorize White Blood Cells Other Tests Hematology Hematology: ◦ Sed Rate- Erytrocyte Sedimentation Rate (ESR) ◦ Sedimentation rate rbc in a given time. ◦ RBCs fall to the bottom in a vertical tube ◦ Non-specific marker of inflammation--- similar C-Reactive Protein Coagulation ◦ Clotting factors : PT/ PTT/ TT/ Fibrinogen ◦ Fibrinolysis – D-Dimers Reticulocytes ◦ They are young RBC / contains RNA– continues to synthesize Hgb after to lose nucleus. ◦ For 24 hours after release of bone marrow. ◦ Reports the ability of RBC production by the bone marrow ◦ Important in blood loss or hemolysis Methodology / Procedure: ◦ Vital Staining (Supravital): Methylene Blue–– stains RNA– manual / automated ◦ Nucleic Acid Dye--- absorbed by living cells ◦ Wright Stain - Polychromasia ◦ Absolute number / percent % / Immature fraction (IRF) Hematology: Other Tests Advanced Hematology Studies: ◦ Bone Marrow Aspirates / Biopsies ◦ Cytochemical Stains: ◦ Myeloperoxidase, Sudan Black. Specific / non-specific esterase/ ◦ Periodic Acid Shift (PAS)/ Alkaline phosphatase/ Tartrate Resistance Acid Phosphatase. ◦ Flow Cytometry– Immunophenotype ◦ Cytogenetics Analysis ◦ Molecular Diagnostic Assays Morphology Blood Smear Hematology Cell morphology The Importance of Cell Examination ◦ Estimates hematologic functions ◦ Presence of disease ◦ Help in hematological diagnosis ◦ Suggest additional tests ◦ Determining the percentage of each of the cell types present is an important skill to master.  Erythrocytes Biconcave disc No nucleus Cell morphology RBC Erythrocytes It is evaluated in stained paving area where they are well distributed rbc. ◦ Do not overlap the RBCs. ◦ RBCs consist of a plasma membrane surrounding a protein solution, primarily hemoglobin and electrolytes. ◦ Normal, mature RBC is a biconcave disc ◦ Average diameter 7 - 8 um ◦ Thickness of 1.5 – 2.5 um  Cell Morphology Erythrocytes RBC After the Blood Smear is dyed with Wright's Stain ◦ Circular cell with distinct and soft margins and has an opaque pink tone. ◦ Fairly uniform in size and round shape. ◦ It has a small area of central pallor and no nucleus or inclusions. ◦ The central portion: ◦ Is Slimmer ◦ The intensity of the dye is less than the margins  Erythrocytes Extended Distributed RBC Wright's Stain Morphology: Anisocytosis – sizes Poikilocytosis – forms  Blood Smear Newborn Normal View nucleated RBCs Polychromasia: blue-gray RBC (reticulocytes) Elevated Hemoglobin & HCT 18 – 22 g/dl Cell Morphology Platelets (thrombocytes) ◦ Diameter from 1 – 4 um and vary in shape ◦ Number of platelets: 7 – 15 plts/field oil immersion ◦ An estimate of the number of platelets in 10 oil immersion fields is made. ◦ The average number of platelets seen is calculated and multiplied by 20,000/mm3. Wright's Stain: A platelet contains purple red granules in a small bluish cytoplasm. It has no nucleus. It contains molecules necessary for hemostasis Able to adhere, aggregate and provide surface for coagulation reaction.  Platelets 7 – 15 / Oil Field Estimated PLTs: The average number of platelets seen 100X Microscope objective is calculated and multiplied by 20,000/mm3. Macroplatelets Platelets Clumped platelets Coagulated sample!! “Satellite phenomenon” Cell Morphology White Blood Cells(WBC) ◦ Neutrophils ◦ Segmented ◦ Bands ◦ Eosinophils ◦ Basophils ◦ Lymphocytes ◦ Monocytes Estimated WBCs: 40X Microscope Objective Add 10 Fields -- average Multiply by 2,000. Cell morphology WBC Normal Values in Adults WBC % Absolute number / mm3 Neutrophil bands 2-6 100 - 650 Segmented neutrophils 50 - 70 2,400 – 7,500 Eosinophiles 0-4 0 - 450 Basophiles 0-2 0 - 200 Lymphocytes 20 - 44 1,200 – 3,400 Monocytes 2–9 100 – 900  Physiological Variations in Children Cells Newborns 1st year Neutrophils 52% 28% Bands 9% 3% Eosinophils 2% 3% Basophils 0% 0% Lymphocytes 31% 61% Monocytes 6% 6%  Leukocytes Wright's Stain Cells are evaluated for abnormalities in the nucleus or cytoplasm Immature cells of any type in peripheral blood is abnormal Cell morphology Neutrophils (segmented, filamented, polymorphonuclear) ◦ Segmented refers to multi-lobed nucleus ◦ 50 – 70% mature granulocytes ◦ The nucleus is divided between two to five lobes ◦ contains narrow segment (filament) connecting the lobes. ◦ 6% one lobe (band) ◦ 35% two (2) lobes ◦ 41% three (3) lobes ◦ 17% four lobes ◦ 2% five lobes Segmented Normal Cell Morphology Neutrophils (segmented, filamented, polymorphonuclear) Segmentation of the nucleus allows it to pass through an opening in the endothelial lining of cells in the capillaries. (Infection area). ◦ They migrate into the tissue as the first line of defense against infection ◦ The chromatin of the nucleus is: ◦ Highly clustered ◦ Gross ◦ Pycnotic ◦ Dyes purple-red  Neutrophils Hyper-segmented Neutrophil Hypo segmented Cell Morphology Neutrophils (segmented, filamentous, polymorphonuclear) ◦ The cytoplasm is pale pink or "lavender“ ◦ Secondary granules ◦ They are thin, numerous, and evenly distributed. ◦ They dye pink or a neutral color. ◦ Contains bactericidal substances: ◦ They are lysosomes that contain alkaline phosphatase. ◦ Size: double to RBC ◦ Neutrophilia (Bacterial infection sign) ◦ Neutropenia (Many causes, but the most common use of medication or viral infection) ◦ It plays a key role in inflammation and phagocytosis Cell morphology Neutrophil – Band: 2 - 6% ◦ Less mature cell – U-shaped nucleus the S/form of horseshoes. ◦ The ends are parallel in appreciable distance ◦ It has no separate nucleus in lobes connected with filaments. ◦ No segmented nucleus. ◦ There is usually a dark pycnotic mass at each pole where the lobe is meant to form. ◦ The secondary granules are small, evenly distributed, tinge pink, and contain alkaline phosphatase. ◦ Occasionally they may have dark primary granules. ◦ Shift to the Left Neutrophils Bands Cell Comparison Segmented Band Metamyelocyte  Cell Morphology Eosinophils 0 – 4% ◦ Large, round cells, easy to recognize ◦ With shrink granules which has an affinity for the acid stain eosin. ◦ Red-Orange Stain – Wright's Stain ◦ The crystalloid core of the nucleus is composed of a Major Basic Protein (MBP), which binds to the acidic aniline dye ◦ Pellets are full of proteins wrapped in the regulation of the immune system. ◦ Spherical granules uniform in size and well distributed ◦ They can be recognized in unstained preparations with light and phase microscopy. Cell morphology Eosinophils Normal peripheral blood: 0 – 4% Slightly larger than neutrophils Nucleus in band or two lobes with condensed chromatin. Rare to find eosinophils with three lobes. Diurnal and nocturnal variation- with an increase in eosinophils circulating at night. Eosinophilia -- Parasitic Infections or Allergy Response Eosinophils Band Two lobes  Cell Morphology Basophils 0 – 2% ◦ Cells with large, abundant granules, blue-violet or purple-black. ◦ Granules filled with Histamine and other proteins. Easy to recognize ◦ The granules darken more than the core in the Wright's stain. ◦ Granules are coarse and vary in size: 0.2 – 1.0 um ◦ They are not evenly distributed. ◦ They vary in number, shape, and color. ◦ Granules are less numerous than eosinophils. ◦ Basophilia– Rare and associated hematological diseases  Basophil Basophilia rare and associated hematological diseases  Cell morphology Basophils Granules have an affinity for blue or basic thiazine dyes. ◦ They are soluble in water– Careful staining process. ◦ If there is poor fixation, during staining, the center of the granule may disappear or the entire granule may be washed away, leaving small areas of the cytoplasm colorless. ◦ Shows diurnal variation similar to eosinophils. ◦ Increases at night. Basophils Water-soluble granules Cell Morphology Lymphocytes 20 – 44% ◦ They are the second most numerous cells in the blood ◦ Responsible for the Immune System (Humoral/Cellular) ◦ Most are small cells; They vary between 7 – 10 um. ◦ They are usually round, with smooth margins and little cytoplasm. ◦ Rare to see in the shape of a spindle (elongated, ellipsoid) ◦ A little bigger than rbc. ◦ They also come in intermediate and large sizes. ◦ Cytoplasm dented by neighboring RBCs. ◦ Lymphocytosis (associated with viral infections) ◦ Lymphopenia (associated with therapeutic drugs or immunodeficiency)  Lymphocyte Usually Measures: 8 –10 um Cell Morphology Lymphocytes In Wright's stain, the color of the cytoplasm is blue, and they vary in intensity. The intensity of the blue is greatest at the periphery of the cell. Clearest near the Golgi area which is adjacent to the core. Cytoplasm is clear, not cloudy. Most don't have granules. In large lymphocytes, few well-defined granules can be seen that vary in size, are not evenly distributed, and are easily counted. They are purple-red – erroneously called azurophils Cell Morphology Lymphocytes Round or lightly toothed Nucleus. ◦ N:C-4:1 to 2:1 ◦ Chromatin is lumpy or clumped. ◦ It stains dark purple with slightly bluish-purple areas between the chromatin aggregates. ◦ They may have a nucleolus present, but it is not visible because of the darkness of the staining. ◦ These cells can grow and replicate.  Lymphocytes Normally seen in infants  Lymphocyte Azurophilic granules Cell Morphology Monocytes Larger cells measuring 12 – 18um. ◦ Monocytosis– Associated with Chronic Infections or Inflammation ◦ Cytoplasm abundant relative to its N:C- 1:1 nucleus; 2:1 ◦ In Wright Stain cytoplasm stains gray-blue. ◦ It has numerous fine, small, reddish, purple granules, evenly distributed. ◦ Cytoplasm with cloudy frosted glass appearance. ◦ It also has prominent granules. ◦ If it is observed not granular - suggests fast turnover ◦ Digestive vacuoles are observed. ◦ It can be seen engulfing rbc, nucleus, bacteria, etc.  Monocyte Immature macrophages: passing through the circulation from their point of origin (bone marrow) to target tissue. Cell Morphology Monocytes Nucleus often in the form of kidneys. Deeply bent or serrated, occasionally lobular. Distinctive features: appearance of convolutions in the nucleus Nucleus like a delicate lattice-like lace with fine strands interspersed with small clustered chromatins. The shape of the cell is variable and can show with pseudopods Monocytes circulate between 8 hours and 3 days before reaching the tissue and transforming into macrophages. Macrophages – the most abundant cells in the body All body cavities – motile and non-motil Function: phagocytosis, presenting Antigns to lymphocytes to mount an immune response. MONOCYTE Monocytes  Cell Morphology Large lymphocytes Monocytes ◦ Clustered core 12 – 15 um ◦ Nucleus as “Net" as lace 12 – 18 um ◦ Condensed core on the periphery ◦ Nucleus-Convolutions Like the Brain ◦ Purple red granules stand out and can be ◦ Prominent bluish-red granules and cannot be easily counted. listed because they are together with fine and ◦ Cytoplasm clear, background not granular. small granules. ◦ Cell indented by neighboring rbc. ◦ Cytoplasm as emerald glass smoothed by fine and small granules. ◦ "ground-glass appearance" ◦ Pseudopods among the cells Monocyte vs lymphocytes Great Challenge to differentiate Reactive Lymphocytes and Monocytes Monocyte vs lymphocytes A- MONOCYTE B- LYMPHOCYTE Cell Comparison Reactive lymphocyte Monocyte  CBC Blood Smear Analysis Parameter Correlation RBC/ Hgb/ Hct Multiplicate by 3 +/- 3 Estimated WBC Count Microscope Obj 40X Add 10 campos -- average Multiply by 2,000. Estimated Platelet Count Microscope Obj 100X oil Adds 10 campos -- average Multiply by 20,000. Reports: adequate, increased, decreased Hematology References Ranges Assay Unit Reference intervals RBC, male x 106 /uL ( X 1012/L) 4-6 RBC, Female x 106 /uL ( X 1012/L) 3–5 Hgb, male g/dl (g/L) 14 – 18 Hgb, female g/dl (g/L) 12 – 16 Hct, male % (L/L) 40 - 54 HCT, female % (L/L) 36 – 47 MCV fL 80 - 100 MCH pg 26 – 34 MCHC g/dl 32 - 36 RDW-CV % 11 – 14 RETICS % 0.5 – 2.5 Hematology References Ranges Assay Unit Reference intervals WBC x 103 /uL ( x 109/L) 4 – 10 50 – 70 ( 2 - 7) Neutrophils % (x 103 /uL ) Bands included (2 – 6%) Lymphocytes % (x 103 /uL ) 20 – 40 (1– 4 ) Monocytes % (x 103 /uL ) 2 – 10 (0.1– 1.0) Eosinophils % (x 103 /uL ) 1 – 4 (0- 0.4) Basophils % (x 103 /uL ) 0 – 2 (0.2 – 2.0) PLTS x 103 /uL ( x 109/L) 150 – 450 MPV fL 7 – 12