W8 Hematology Part 1.pdf

1st SEMESTER ‘24-’25 | MIDTERM Lecturer: Prof. Rene Jesus Alfredo Dinglasan WEEK 8: HEMATOLOGY PT 1 ○ According to Turgeon, EDTA is the preferred anticoagulant for Topic Outline:...

1st SEMESTER ‘24-’25 | MIDTERM Lecturer: Prof. Rene Jesus Alfredo Dinglasan WEEK 8: HEMATOLOGY PT 1 ○ According to Turgeon, EDTA is the preferred anticoagulant for Topic Outline: mean platelet volume (MPV), but it causes platelets to swell A. Anticoagulants E. Erythropoiesis (approximately a 20% increase in MPV during the first hour— B. Order of Draw in Venipuncture F. RBC Stages of Maturation after this, the size is stable for at least 12 hours). Therefore, C. Errors Encountered in some G. Red Blood Cell Anomalies H. Hemoglobin MPV measurements should be based on EDTA specimens that Hematology Tests D. Critical Values that Require I. Hematocrit Determination are between 1 and 4 hours old. Immediate Communications ○ Measure of the average volume of platelets in a sample ○ Analogous to the erythrocytic MCV ANTICOAGULANTS ○ In healthy patients, there is an inverse relationship between 1 EDTA (ETHYLENEDIAMINETETRAACETIC ACID) platelet count and size: volume increases as platelet count The usual anticoagulant used in Hematology decreases. Usually as K2EDTA EDTA is recommended by the ICSH and CLSI as the anticoagulant Most commonly used blood collection tube in the Hematology of choice for blood cell counting and sizing. section Proper ratio of EDTA to whole blood is important because some test results will be altered if the ratio is incorrect. Excessive EDTA Found In Lavender/purple top tube produces shrinkage of erythrocytes, thus affecting tests such as No. of Inversions 8x manually performed packed cell volume (microhematocrit). For routine hematology determination Use INSUFFICIENT EDTA EXCESSIVE EDTA ○ The correct tube to use for CBC Cause: overfilled Cause: underfilled/“short draw” 1.5 mg/mL of blood — In order to prevent Optimum tube tubes coagulation in every milliliter of blood, you Concentration Effect: presence of Effect: will need this amount of EDTA. clots ○ ↓ Hct (Hematocrit) Chelation of Calcium ○ ↓ ESR (Erythrocyte ○ Chelation is the formation of a ring- Sedimentation Rate) shaped molecular complex in which a ○ Degenerative changes in WBC metal ion is covalently bound. ○ ↑ MCHC (Mean Cell In other books, the action of EDTA is Hemoglobin Concentration) Action referred to as the 'binding of calcium' ○ ↑ Platelet count instead of chelation. However, both terms can be used, as chelation is just a specific Note: way of binding ions. On the bottom of the EDTA tube, the volume of blood is indicated It forms an insoluble calcium salt that such as '4mL'. The volume of the blood must be exactly 4mL, not prevents blood coagulation. 5mL, because the blood will overpower the anticoagulant, causing clots. IMPORTANT REMINDERS IN USING EDTA: Clots can damage machines and may alter CBC parameters of Blood specimens for CBC must be analyzed within 6 hours of the patient. collection (if stored at room temperature), and 24 hours (if stored EDTA tubes are not all the same; some are 4 mL, 5 mL, or 7 mL. at 4°C/refrigerator) to minimize false results. The use of the ETS (Evacuated Tube System) is better than ○ We place it inside the refrigerator so we can reuse it for up to syringes, since the tubes are filled using their own vacuum, 24 hours. Sometimes, doctors want to recheck CBC values, so ensuring that the exact amount of blood is extracted. we need to preserve it for 24 hours. This way, if a doctor calls the laboratory to retest specific CBC parameters, we can still BOARD EXAM QUESTIONS! provide the results. What is the preferred anticoagulant for platelet count? Blood smears should be made within 3 hours of collection, not Answer: EDTA, BECAUSE IT PREVENTS PLATELET beyond that, because if it’s longer, the blood cells may be destroyed AGGREGATION. or distorted. EDTA prevents platelet aggregation; therefore, it is the preferred What is the color of the most commonly used tube in the anticoagulant for platelet counts. Hematology Section? Mean Platelet Volume (MPV) Answer: PURPLE/LAVENDER TOP Transcribers: Ambas, Angcao, Apostol, Austero, Balines, Cabalang, Calpatura, Camingawan, Carpio, Castillones, De Leon, Debaja, 1 of 26 Deinla, Delos Santos, Donato, Etcobanez, Gayona, Gendrano, Gotico, Ignacio, Lalas, Lu, Mangulad, Mercado, Miranda, Moreno, Nasinopa, Pacleb, Paculba, Pagulayan, Perez, Sapla, Sabareza, Silvestre, Tan, Uy, Ventura, Villanueva, Yatco  What is the correct time frame for performing MPV? Osmotic Fragility Test (OFT) Blood Gas Studies Answer: EDTA SPECIMENS THAT ARE BETWEEN 1-4 HOURS OLD Optimum 15-20 Units per mL of blood Concentration It binds antithrombin. COMPLETE BLOOD COUNT (CBC) Heparin is not actually an anticoagulant. It Also called HEMOGRAM has no anticoagulant power on its own. It Performed on automated hematology profiling instruments and Action derives its anticoagulant activity from includes: antithrombin. Once bound with ○ RBC Parameters: antithrombin, it can now inhibit thrombin RBC Count and Factor Xa. Hemoglobin Hematocrit THREE (3) HEPARIN FORMULATIONS RBC Indices 1 AMMONIUM HEPARIN Red Cell Distribution Width (RDW) Must NOT be used for ammonia level determination Reticulocyte Count ○ WBC Parameters 2 SODIUM HEPARIN WBC Count Must NOT be used for sodium level determination or for electrolyte NEUT count: % and absolute panel LYMPH count: % and absolute 3 LITHIUM HEPARIN MONO count: % and absolute EO and BASO counts: % and absolute Must NOT be used for lithium level determination ○ Platelet Parameters Causes the least interference in chemistry testing Platelet Count Most widely used anticoagulant for plasma and whole blood Mean Platelet Volume (MPV) chemistry tests IMPORTANT REMINDERS IN USING HEPARIN: POSSIBLE CAUSES OF CLOTTED BLOOD SPECIMENS Heparin causes cellular clumping (especially of platelets), which Difficult phlebotomy leads to pseudoleukocytosis (falsely INCREASED WBC count) Blood sample was not inverted in the correct number of times and pseudothrombocytopenia (falsely DECREASED platelet Expired blood collection tube count) on some hematology analyzers. Overfilled tube ○ It is not used in CBC. RELATED TERMS Not to be used for blood smear preparation because it causes DELTA CHECK: a process in which a current test result is morphologic distortion of platelets and leukocytes compared with the result of the same test from the previous Causes bluish coloration of the background on blood smears stained specimen from the same patient with a Romanowsky stain because of its pH ○ Delta check method of QC utilizes the patient's own data to Not to be used for coagulation studies because it inhibits thrombin monitor population values. Flow cytometry and OFT are both performed in Hematology. REFLEX TEST: a test being ordered automatically based on the Plasma chemistry determinations and blood gas studies are done results of prior tests or preset parameters in the Clinical Chemistry section. PARAMETER: any numerical value that depicts an entire population Automations related to flow cytometry were also asked about 2 HEPARIN during the board exam. This is the anticoagulant of choice for OFT and blood gas studies. 3 3.2% SODIUM CITRATE Heparin is not usually used in the Hematology section. 3.2% Sodium Citrate is found in light blue top tubes and is used ○ EDTA is the most common, especially for Complete Blood for coagulation tests. Count (CBC). 3.8% Sodium Citrate is found in black top tubes and is used for There are some hematologic tests that use heparin as an ESR. anticoagulant, such as OFT. Found In Light blue top Found In Green top tube 3-4x No. of Inversions 8x ○ Forceful mixing or an excessive Flow Cytometry No. of Inversions Use number of inversions can activate Plasma Chemistry Determinations platelets and shorten clotting times 2 of 26  ○ Proper inversion shall also be Lavender/Purple/Pink EDTA Tube 8 observed. The manufacturer's Top guidelines must be followed. Sodium Fluoride 8 Gray Top Remember, it is inversion and not shaking. Note: Use For coagulation tests PINK TOP TUBE is the appropriate alternative if there are no Optimum The critical ratio between anticoagulant lavender tops available. Concentration and blood is 1:9. “The placement of tubes not listed here should take into Chelation of Calcium consideration the potential for their additive to alter results Action ○ It is the same thing performed by obtained from the next tube if carryover were to occur. EDTA. Plastic serum tubes containing a clot activator may cause interference in coagulation testing. Only blood culture tubes, glass BOARD EXAM QUESTIONS! nonadditive serum tubes, or plastic serum tubes without a clot What is the critical ratio between anticoagulant and blood? activator may be collected before the coagulation tube.” Answer: 1:9; 1 PART ANTICOAGULANT TO 9 PARTS BLOOD ○ Source:https://clsi.org/about/blog/order-of-blood-draw- tubes-and-additives/ Note: The board exam is given by licensed MedTechs who are not ERRORS ENCOUNTERED IN SOME HEMATOLOGY TESTS connected to any MedTech schools; thus, it focuses on PARAMETERS TYPE OF laboratory procedures, and we must understand the reasons and AFFECTED ERROR PROBABLE REASON/S methods behind these procedures. Falsely WBC > 100,000/μL, large(giant) high platelets, cryoglobulin, cryofibrinogen REMINDERS: RBC count autoagglutination, cold agglutinins, Collect the blood specimen of your patient using the correct order of Falsely hemolysis (in vitro), microcytosis, draw ow schistocytes, clotting ○ Purpose: to avoid possible test errors because of cross- autoagglutination, high WBC contamination from tube additives (>50,000/μL), cold agglutinins, old Invert each tube containing additive immediately after Falsely specimen, hyperosmolar state high collection. Mean Cell Volume (uncontrolled diabetes mellitus), Label the blood collection tubes appropriately. The minimal amount (MCV) reduced red cell deformability of information that should be on each tube is as follows: cryoglobulin, cryofibrinogen, giant Falsely ○ Patient’s full name platelets, hemolysis (in vitro), swollen high ○ Patient’s unique identification number red cells ○ Date and time (military time) of collection lipemia, icterus, chylomicrons, high Falsely WBC (>50,000/μL), spuriously high ○ Phlebotomist’s initials or code number Mean Cell high Hemoglobin Hb, spuriously low RBC ○ Compare the labeled tube with the patient’s identification bracelet or have the patient confirm that the information on the (MCH) Falsely low spuriously low Hb, falsely high RBC blood collection tube is correct (whenever possible). cold agglutinins, autoagglutination, clotting, hemolysis (in vitro), ORDER OF DRAW IN VENIPUNCTURE Falsely Mean Cell hemolysis (in vivo), spuriously high high Order of Draw Mnemonic: BC, SHES Hemoglobin Hb, falsely low Hct The order of draw is very important to prevent contamination of the Concentration Falsely high WBC (>50,000/μL), falsely low tubes. (MCHC) low Hb, spuriously high Hct BLOOD NO. OF lipemia, icterus, chylomicrons, lysis- COLOR COLLECTION TUBE INVERSIONS resistant RBCs with abnormal Hbs, Blood Culture Tubes 8 Yellow Top WBC > 20,000/μL (>20 x 109 /L), platelet count (>700 x 109 /L), Citrated Tube 3-4 Light Blue Top Falsely Hemoglobin parenteral nutrition, hypergamma- Red Top (glass) (non- high 0 globulinemia, cryoglobulin, additive) cryofibrinogen, hemolysis (in vitro), Serum Tube Red Top (plastic) heparin, hyperbilirubinemia 5 (has clot activator) Falsely low clotting Heparinized Tube 8 Green Top 3 of 26  Dehydration, hemoconcen- Any critical value obtained must be documented and referred Falsely tration,insufficient centrifugation, immediately to the pathologist. A preliminary report may be issued high buffy coat inclusion, hyponatremia, followed by a final report after review by the pathologist. plasma trapping Hematocrit TEST CRITICAL VALUES hemolysis (in vitro), improper sealing (micro- of the capillary tube, increased ≤ 21% or ≤ 0.21 hematocrit) Falsely Hematocrit anticoagu- lant concentration, ≥ 65% or ≥ 0.65 low introduction of excess tissue fluid, ≤ 70 g/L hypernatremia Hemoglobin ≥ 200 g/L cryoglobulin, cryofibrinogen, giant Falsely platelets, high WBC (>50,000/μL), Hematocrit high hyperglycemia (>600 mg/dL) Reticulocyte > 20% (automated) Falsely autoagglutination, clotting, hemolysis low (in vitro), microcytic red cells < 2,000/uL on a new patient or a 1,000 difference from lysis-resistant RBCs with abnormal WBC count previous, if less than 4,000/uL Hbs, nucleated RBCs, > 50,000/uL on a new patient megakarycyte fragments, Neutrophilic phagocytosis of microorganisms (indicating Falsely micromegakaryoblasts, platelet the presence of systemic infection) high clumps,giant platelets, cryoglobulins, Shows abnormal leukemoid reaction WBC count cryofibrinogen, heparin, monoclonal Schistocytes (may indicate hemolytic condition) Blood Smear proteins Sickle cells leukemia (especially with Blast forms (if never or not recently reported in the Falsely chemotherapy), leukoagglutination, patient) low clotting, smudge cells Presence of intracellular organisms neutrophil aggregation, neutrophil < 20,000/uL and NOT previously reported Falsely Platelets Neutrophils with hemosiderin granules (were > 1 million/uL low counted as eosinophils) Prothrombin > 40 seconds nucleated red cells, giant platelets, Time Lymphocyte Falsely platelet clumps, malarial parasites, high hypolobated neutrophils BOARD EXAM QUESTIONS! large reactive lymphocytes, What is the value of the WBC count that can cause a falsely high Monocyte Falsely lymphoblasts, lymphoma cells, hemoglobin result? high immature granulocytes neutrophil with hemosiderin granules Answer: WBC COUNT: > 20, 000 /UL OR > 20 X 10^9 /L OR HIGH Falsely Eosinophils (counted as eosinophils), red cells PLATELET COUNT: > 700, 000/ UL OR > 700 X 10^9 /L high with malarial pigments leukemia (especially with What is the level of prothrombin time considered critical for a chemotherapy),fragmented red cells patient or life-threatening values? (in micro- angiopathic hemolysis), Falsely Answer: > 40 SECONDS microorganisms,cryoglobulins, high cryofibrinogen, hemolysis (in vitro Platelet count and in vivo), microcytic red cells, red ERYTHROPOIESIS cell inclusions, white cell fragments The process of red blood cell (RBC) formation which takes platelet clumps, old specimen, partial place inside the bone marrow. Falsely clotting, giant platelets, platelet The CFU-GEMM gives rise to the earliest identifiable colony of low satellitism, cold agglutinins, clotting, RBCs, called the burst forming unit-erythroid (BFU-E). The BFU-E heparin produces a large multiclustered colony that resembles a cluster of MPV (Mean old specimen, determining the MPV Falsely high grapes containing brightly colored hemoglobin. Platelet Volume) too early in an EDTA specimen RELATED TERMS CRITICAL VALUES THAT REQUIRE IMMEDIATE COMMUNICATIONS 1 ERYTHRON Critical values must be determined by the laboratory in consultation The total mass of RBCs circulating in the peripheral blood and with the users. the bone marrow RBC precursors. When a critical value is obtained, the physician of the patient or the It primarily refers to the total population of RBC in the blood and responsible healthcare worker must be informed by any means of bone marrow. It includes all erythrocytes regardless of where they communication. are. 4 of 26  ○ Erythron is the name given to the collection of all stages of What is considered as the birthplace of the RBC? erythrocytes throughout the body: the developing precursors in the bone marrow and the circulating erythrocytes in the Answer: Bone Marrow peripheral blood and vascular spaces within organs, such as What is the average life span of RBC after they are released from the spleen. the bone marrow? 2 RBC MASS Answer: 120 days The erythrocytes in the circulation. It refers to the total population of RBC in the circulating blood ONLY, excluding those in the bone marrow 6 PROGENITOR CELL The erythron is distinguished from the RBC mass. An immature hematopoietic cell that is committed to a cell line but The erythron is the entirety of erythroid cells in the body, whereas CAN’T BE IDENTIFIED morphologically through a light the RBC mass refers only to the cells in circulation. microscope. Example: 3 ERYTHROKINETICS ○ BFU-E (Burst Forming Unit - Erythroid) A term that describes the dynamics of RBC creation and ○ CFU-E (Colony Forming Unit - Erythroid) destruction. 7 PRECURSOR CELL 4 INEFFECTIVE ERYTHROPOIESIS An immature hematopoietic cell that is morphologically The production of defective erythroid precursor cells. IDENTIFIABLE as belonging to a given cell line. These defective RBC precursors often undergo apoptosis Example: (programmed cell death) in the bone marrow; they have a chance to ○ Rubriblast mature to the reticulocyte stage. ○ Prorubricyte Since they are already dead inside the bone marrow, it will eventually develop anemia. 8 CD71 CONDITIONS IN INEFFECTIVE ERYTHROPOIESIS The earliest marker of erythroid differentiation MACROCYTIC, MICROCYTIC, HYPOCHROMIC Serves as a transferrin receptor for transferrin coming from the NORMOCHROMIC plasma. ○ Receptor is responsible for receiving Vitamin B12 deficiency Thalassemia ○ Transferrin is the transport protein of iron in the blood Folate deficiency Sideroblastic anemia Note: 5 INSUFFICIENT ERYTHROPOIESIS Albumin is the transport protein of Bilirubin 1 (unconjugated There is a decrease in the number of RBC precursors in the bilirubin) bone marrow resulting in the decrease in RBC production. Ceruloplasmin is the transport protein of copper in the blood Consists of 3 types of anemia: Von Willebrand Factor (VWF) is the transport protein of Factor ○ Macrocytic, normochromic VIII in the plasma. ○ Normocytic, normochromic ○ Microcytic, hypochromic HORMONES RELATED TO ERYTHROPOIESIS CONDITIONS IN INSUFFICIENT ERYTHROPOIESIS 1 ERYTHROPOIETIN (EPO) NORMOCYTIC, Chief stimulatory cytokine for RBCs MICROCYTIC, HYPOCHROMIC Major hormone that stimulates the production of erythrocytes NORMOCHROMIC Renal Disease Thermostable, nondialyzable, glycoprotein hormone Iron deficiency First human hematopoietic growth factor to be identified Acute leukemia Produced primarily by the kidneys (80-90%) Notes: ○ 10-15% of EPO production occurs in the liver which is the Necrosis is described as the pathological cell death. primary source of EPO in the unborn Anemia is not a disease, but a manifestation of an underlying Primary cell source: Peritubular interstitial cell (kidneys) disease. ○ Normally, EPO is released from the kidney into the blood in response to hypoxia (too little tissue oxygen); blood levels of BOARD EXAM QUESTION! EPO are inversely related to tissue oxygenation Primary target cells: BFU-E & CFU-E (progenitor cells) What is considered as the graveyard of the RBC? Answer: Spleen 5 of 26  3 MAJOR EFFECTS: Notes: 1. Allowing early release of reticulocytes from the bone marrow Testosterone in the body of males increases erythropoiesis. 2. Reducing the time needed for cells to mature in the bone Whereas, estrogen in the body of female inhibits erythropoiesis marrow DIRECT stimulation of erythropoiesis: 3. Preventing apoptotic cell death ○ Erythropoietin, growth hormone, testosterone, and prolactin ○ Apoptosis rescue is the major way in which EPO INDIRECT stimulation of erythropoiesis: increases RBC mass ○ Estrogen Some of the current/potential therapeutic applications: ○ Anemia of chronic renal disease Why are males RBC count, Hgb, and Hct higher than ○ Autologous predonation blood collection females? ○ Anemia in HIV infection to permit use of zidovudine (AZT) ○ Females generally have a lower red blood cell (RBC) count “BLOOD DOPING” compared to males, not primarily because of menstruation, ○ Some athletes illegally use EPO injections to increase the but due to the hormone ESTROGEN. Estrogen inhibits oxygen-carrying capacity of their blood (to enhance endurance erythropoiesis, which in turn limits the stimulation of RBC and stamina) (can lead to deadly arterial and venous production. thrombosis) ○ In contrast, males typically have higher RBC count, hematocrit (Hct), and hemoglobin (Hgb) levels because the BOARD EXAM QUESTION hormone TESTOSTERONE stimulates erythropoiesis and Which of the following hormones produced by the kidneys is for enhances RBC production RBC production? ○ While both sexes produce testosterone and estrogen, the A. Erythropoiesis balance of these hormones differs, with males having higher B. Erythropoietin testosterone levels and females having higher estrogen Answer: B. Erythropoietin levels IN TRANSGENDER WOMEN: 2 GROWTH HORMONE ○ Before Transitioning (when presenting as male): higher Produced by the pituitary gland levels of RBC count, hemoglobin (Hgb), and hematocrit ○ Specifically, GH originates from the anterior pituitary gland (Hct) due to the influence of testosterone. Able to stimulate Erythropoiesis DIRECTLY ○ After Transitioning (when presenting as female): These ○ It goes to the bone marrow itself to produce more RBCs levels decrease as they undergo hormone replacement (Growth hormone → bone marrow → RBC production) therapy, which involves taking estrogen to develop more traditionally feminine characteristics. 3 TESTOSTERONE ○ Effects of Estrogen: Produced by the testes Bigger chests Able to stimulate Erythropoiesis INDIRECTLY Wider hips ○ Goes to the kidneys to be stimulated to produce more Smaller waists Erythropoietin (EPO), which then acts on the bone marrow to Lowering RBC count, Hgb, and Hct stimulate RBC production. (Testosterone → kidneys → EPO ○ Transgender women have low HCT, Hgb, and RBC count → bone marrow → RBC production) because of the hormones and even though they produce these, they still do not have MENSTRUATION. Note: DIRECT way = Bone marrow RBC STAGES OF MATURATION INDIRECT way = Kidneys 2 COMMON ERYTHROID PROGENITOR CELLS 4 PROLACTIN (PRL) Burst forming unit-erythroid BFU-E Produced by the pituitary gland The EARLIEST committed progenitor Able to stimulate Erythropoiesis DIRECTLY CFU-E Colony forming unit-erythroid ○ Goes to the bone marrow itself to produce more RBCs BFU-E to CFU-E: 1week CFU-E to Rubriblast: 1 week or 7 days 5 ESTROGEN BFU-E to Mature erythrocyte: 18-21 days or 3 Produced by the ovaries Duration weeks Inhibits Erythropoiesis INDIRECTLY ○ Out of the 18 to 21 days it takes for erythroid ○ Goes to the kidneys to inhibit the synthesis of Erythropoietin precursors to develop in the bone marrow, Has reverse effects from Testosterone they become recognizable around day 6. 6 of 26  According to Turgeon, the maturation time from myelocyte to AKA Nucleated RBC (NRBC), Pyknotic Metarubricyte erythroblast, Acidophilic normoblast metamyelocyte is 4.3 days. The prefix “Meta” means “after” 3 NOMENCLATURES OF RBC STAGES ○ Therefore, metarubricyte is the cell that follows rubricyte. RUBRIBLASTIC NORMOBLASTIC ERYTHROBLASTIC Reticulocyte 5th precursor Rubriblast Pronormoblast Proerythroblast Basophilic Normoblast Basophilic erythroblast or Mature 6th stage/ Last stage Prorubricyte or Early Erythroblast Erythrocyte NOT a precursor Early Normoblast Polychromatic Polychromatophilic BOARD EXAM QUESTION! Rubricyte Normoblast or erythroblast or Intermediate Intermediate Erythroblast Which of the following is the last stage capable of mitosis? Normoblast a. First precursor Orthochromatic Orthochromatic b. Second precursor Metarubricyte Normoblast or erythroblast or c. Third precursor Late Normoblast Late Erythroblast d. Fourth precursor Reticulocyte Reticulocyte Reticulocyte Answer: Third precursor or Rubricyte Mature erythrocyte Mature Erythrocyte Mature Erythrocyte Always remember the correct order of the precursors since sometimes the actual names are not given in exams (Ex. Precursor 1, Precursor 2, Precursor 3) STAINS For reticulocytes: ○ Brilliant Cresyl Blue (BCB) ○ New Methylene Blue (NMB) For reticulocytes found in WRIGHT STAIN : ○ Polychromatophilic Erythrocytes ○ Diffusely Basophilic Erythrocytes RUBRIBLASTIC NOMENCLATURE Consists of 5 erythrocyte precursor (immature) cells and 1 mature erythrocyte, making up a total of 6 stages. STAGE OTHER NAMES Notes: 1st precursor When a reticulocyte is stained with Wright's stain, it should not “Blast” is always considered as the 1st Rubriblast be referred to as a "reticulocyte." Instead, it should be called a precursor polychromatophilic erythrocyte or diffusely basophilic ○ Ex: Monoblast, Lymphoblast, erythrocyte. Myeloblast, Megakaryoblast ○ Supravital stain: Reticulocyte 2nd precursor ○ Wright stain: Polychromatophilic erythrocyte or Diffusely Prorubricyte The prefix “Pro” means “before” basophilic erythrocyte. ○ So, a Prorubricyte is a cell that comes before the rubricyte. “Polychromatic” and “Polychromatophilic” are the same / ○ Ex: Promyelocyte,Promonocyte synonymous. Prolymphocyte,Promegakaryocyte Methylene blue and New methylene blue (NMB) are different Rubricyte 3rd precursor from each other. Methylene blue cannot be used as a substitute 4th precursor for Brilliant Cresyl Blue since they have different components. 7 of 26  QUESTIONS: cytoplasm of the prorubricyte does not show any pink hues. What is the correct name of polychromatophilic erythrocyte in a This is because it is still referred to as a basophilic Wright stained smear? erythroblast, so its cytoplasm stains blue. Answer: POLYCHROMATOPHILIC ERYTHROCYTE OR 3 RUBRICYTE (POLYCHROMATIC NORMOBLAST) DIFFUSELY BASOPHILIC ERYTHROCYTE Size: 11 to 15 μm What is the name of polychromatophilic erythrocyte in a Nucleus: round and smaller, thick nuclear Supravital stained smear? membrane, eccentric nucleus Nucleolus: None Answer: RETICULOCYTE N:C ratio: 1:1 1 RUBRIBLAST (PRONORMOBLAST) Stage Turgeon Rodak’s Size: 12-19 um Rubriblast 8:1 N:C ratio = 4:1 4:1 Prorubricyte 6:1 Nucleus: Round or slightly oval, thin nuclear Rubricyte 1:1 4:1 membrane, central or slightly eccentric Nucleoli: 1 to 2 LAST STAGE capable of mitosis ○ Nucleolus is found in the nucleus of the ○ While this cell can still divide, it is the final stage in red cell cell development that retains this ability. ○ Nucleoli = plural; Nucleolus = singular ○ Succeeding stages, such as metarubricytes, do not have the Cytoplasm: small in amount, dark blue (because of the capacity for mitosis. concentration of ribosomes), homogenous ○ Each rubricyte gives rise to metarubricytes. The cell gives rise to 2 prorubricytes. Cytoplasm: Gray Considered as the EARLIEST recognizable erythroid precursor 1ST STAGE wherein the PINKNESS of the hemoglobin is using the light microscope. manifested in the cytoplasm ○ Precursors are immature hematopoietic cells which can be ○ The gray cytoplasm of the rubricyte is derived from the recognized using light microscopy. combination of the inherent blue color of the precursor’s ○ In contrast, progenitor cells cannot be determined in advance cytoplasm and pink color derived from the hemoglobin. whether they will develop into RBCs or WBCs. ○ This is also why it is called a Polychromatophilic normoblast/erythroblast. 2 PRORUBRICYTE (BASOPHILIC NORMOBLAST) ○ While the prorubricyte begins hemoglobin production, the pink SIZE: 12 to 17 μm coloration is not yet prominent because the amount of N:C ratio: 4:1 hemoglobin is still relatively low. ○ According to Turgeon, the N:C ratio of both ○ As the cell progresses to the rubricyte stage, hemoglobin prorubricyte and rubriblast is 4:1. Rodak’s production increases, making the pink color more noticeable. show different ratios. ○ Therefore, the rubricyte is correctly identified as the first stage Nucleolus: 0 to 1 where the cytoplasm exhibits a pink hue since pinkness of the ○ The prorubricyte is considered as the LAST STAGE with a hemoglobin has manifested. nucleolus. Nucleus: round, thin nuclear membrane, smaller, slightly eccentric Rubricyte Lymphocyte This cell is still capable of mitosis, producing 4 rubricytes after Nucleus Checkerboard Crushed velvet mitosis Cytoplasm Muddy or Sky blue or The most useful criteria for differentiating a prorubricyte from gray Robin egg blue rubriblast are the following characteristics of the prorubricyte: ○ A coarser chromatin pattern 4 METARUBRICYTE ○ The absence of nucleoli Also known as: Cytoplasm: The cytoplasmic color of the prorubricyte has a deeper, ○ Orthochromatic normoblast/ richer blue and appears more abundant than in normoblast because erythroblast of the smaller nucleus. ○ Late normoblast/ erythroblast 1ST STAGE of HEMOGLOBIN SYNTHESIS ○ Nucleated RBC (nRBC) ○ Hemoglobin is an eosinophilic material. It stains pink when ○ Pyknotic erythroblast subjected to staining. ○ Acidophilic normoblast ○ Although the prorubricyte is the 1st stage where hemoglobin Size: 8 to 12 µm synthesis begins and we would expect a pink coloration, the Nucleus: pyknotic (dense mass of degenerated chromatin) 8 of 26  ○ Nucleus is extruded at this stage, and the cell becomes a LAST STAGE of HEMOGLOBIN SYNTHESIS reticulocyte. Young RBCs containing residual RNA (last immature erythrocyte Nucleolus: None stage) Cytoplasm: salmon pink (indicates large quantities of hemoglobin Spends 2 to 3 days in the bone marrow and 1 day in the or complete hemoglobin production) peripheral blood before developing into a mature RBC N:C ratio: 1:2 (low) ○ Under normal conditions, the quantity of reticulocytes in the LAST STAGE wherein the nucleus is present. bone marrow is equal to that of the reticulocytes in the circulating blood. PYRENOCYTE The shape of the reticulocyte is irregular in electron micrographs The extruded enveloped nucleus is called “Pyrenocyte” The formation of new ribosomes ceases with the loss of the nucleus This is usually engulfed by bone marrow macrophages. in the late metarubricyte; however, while RNA is present, protein and ○ In cases wherein small fragments of the nucleus is left behind heme synthesis continues due to the projections being pinched off before the entire If, under the stimulus of erythropoietin, increased numbers of young nucleus is enveloped, the residual fragments seen in RBCs in reticulocytes are prematurely released from the bone marrow the circulation are termed “Howell-Jolly bodies” because of such conditions as acute bleeding, these reticulocytes After this stage, the cells will no longer undergo mitosis. are referred to as stress or shift reticulocytes. Note: “Orthochromatic” Notes: ○ “Ortho” = same Different authors have different opinion regarding the same ○ “Chroma” = color matter of the first stage of hemoglobin synthesis ○ It has the same color of cytoplasm as with the mature erythrocyte In some books, first stage of hemoglobin synthesis happens in Rubriblast QUESTIONS In Cruzada, the first stage is Rubricyte Are Howell-Jolly Bodies nuclear fragments? In Rodaks, the first stage is Prorubricyte Answer: Yes. Since they are nuclear fragments, they are made out of 6 MATURE ERYTHROCYTE DNA. The appropriate stain for DNA materials in the Histopathology section is the Feulgen reaction. Howell Jolly bodies are Feulgen 6th stage of red cell maturation (not a reaction (+) precursor) Size: 6 to 8 um It is stated in Rodaks that we can frequently see Howell-Jolly Nucleus: None bodies in the circulation but how come we do not see Howell- Nucleolus: None Jolly Bodies in normal Peripheral blood smear? Cytoplasm: salmon pink (with a central pallor occupying ⅓ of the cell’s diameter) Answer: These RBC inclusions are typically removed by splenic Shape: Biconcave disk macrophages through the process of pitting once they enter the Thickness: 1.5 to 2.5 um circulation Average life span: 120 days ○ Aging leads to their removal by the spleen. If the patient underwent a splenectomy, will the Howell-Jolly Number of erythrocytes produced from each rubriblast: 8 to 32 bodies increase in values? Normal ratio: ○ RBCs to WBCs = 600:1 Answer: Yes, because there won’t be any macrophages to remove ○ RBCs to Platelets = 15:1 these inclusions. As mentioned, spleen plays a critical role in filtering Adult RBC contains no mitochondria (no protein or Hb synthesis) the blood and removing abnormal cells and inclusions, including Howell-Jolly bodies. SUMMARY Size N:C Nucleolus Cytoplasm Stage (um) ratio 5 RETICULOCYTE Rubriblast 12-19 4:1 1-2 Dark blue Size: 7 to 10 um Prorubricyte 12-17 4:1 0-1 Deep blue Nucleus: None Rubricyte 11-15 1:1 None Gray Nucleolus: none Metarubricyte 8-12 1:2 None Salmon pink Cytoplasm: predominant color is that of Reticulocyte 7-10 - None Salmon pink hemoglobin but with a bluish tinge because Mature 6-8 - None Salmon pink of some residual ribosomes and RNA erythrocyte ○ By the end of this stage, the cell is salmon pink 9 of 26  SUMMARY c Eosinophilia Rubriblast Earliest recognizable erythroid precursor Pertains to PINKNESS of the particular part of the cell that is due to Last stage with a nucleolus the accumulation of more basic components that attract the acid Prorubricyte First stage of hemoglobin synthesis stain stain Eosin. Last stage capable of mitosis As the RBC matures, the eosinophilia of the cytoplasm correlates Rubricyte First stage in which the cytoplasm turns pink with the accumulation of hemoglobin. Metarubricyte Last stage with nucleus RED BLOOD CELL ANOMALIES Reticulocyte Last stage of hemoglobin synthesis Anisocytosis Poikilocytosis CHARACTERISTICS OF MATURATION OF ERYTHROID Anisochromia RBC inclusion bodies PRECURSORS 1 ANISOCYTOSIS As the erythroid precursors mature: Increased number of red cells with variation in size a. The overall diameter of the cell decreases ○ If there are plenty of erythrocyte having a variation in size, there b. The nuclear diameter decreases is anisocytosis c. The chromatin pattern becomes coarser, clumped, and ○ Variation in size: some are 6, some are 7 or 8 um condensed RBCs having variation in size is definitely normal, but d. The nucleoli disappear when there is increased variation in size, it becomes e. The cytoplasm changes from blue to gray-blue to salmon abnormal. pink. Normal RBCs (normocytes): 6 to 8 um in diameter (usually seen when MCV is 80-100 fL) ○ If there are plenty of normocytic cell = MCV will also become normal. To be able to recognize an abnormality or variation in size, it’s important to first remember what is normal. MACROCYTE MICROCYTE Larger than normal RBCs Smaller than normal RBCs (diameter > 8.0 um) (diameter < 6.0 um) Usually seen when the MCV is > Usually seen when the MCV is < 100 fL 80 fL Figure 5.2 (A), Cell diameter decreases and cytoplasm changes from blue to salmon pink. (B), Nuclear Associated with impaired DNA Associated with defective diameter decreases and color changes from purplish-red to a very dark purple-blue. (C), Nuclear chromatin becomes coarser, clumped, and condensed. (D), Composite of changes during the synthesis hemoglobin formation developmental process. RELATED TERMS FOUR WAYS TO DETECT ANISOCYTOSIS a N:C (Nucleus to Cytoplasm) ratio 1 USING THE NUCLEUS OF A SMALL LYMPHOCYTE IN A PBS A morphologic feature used to identify and stage RBC and WBC On the peripheral blood smear, look for a small lymphocyte in precursors order to assess the size of the erythrocyte A visual estimate of what area of the cell is occupied by the nucleus Compare the size of the nucleus of the small lymphocyte with compared with the cytoplasm. the RBCs surrounding it If the areas occupied by the nucleus and cytoplasm are ○ If the RBCs are of similar size with the nucleus of the small approximately equal, the N:C ratio is 1:1 lymphocyte = erythrocytes are normal in diameter or If the nucleus takes up 50% of the area of the cell, the ratio is surrounding it = erythrocytes are small or MICROCYTIC higher (e.g., 3:1 or 3) ○ If the nucleus of the small lymphocyte is smaller than the RBCs surrounding it = erythrocytes are large or MACROCYTIC b Basophilia If the RBCs are normocytic = NO ANISOCYTOSIS Pertains to BLUENESS of the particular part of the cell and is due If the RBCs are either microcytic or macrocytic = WITH to the acidic components that attract the basic stain (ex. Methylene ANISOCYTOSIS blue) Degree of cytoplasmic basophilia correlates with quantity of ribosomal RNA 10 of 26  2 USING THE MCV VALUE INTERPRETATION OF RBC HISTOGRAMS Average volume of individual RBCs a Normocytic RBC population ○ Can be used to assess the size of the erythrocytes or to detect The first step in interpreting an RBC histogram is to locate the peak the presence of anisocytosis of the curve. Reference range: 80-100 fL ○ After that, trace downwards to determine where the peak of the Formula: curve is positioned INTERPRETATION: ○ MCV and RDW are normal. ○ Since RDW is normal, there is no anisocytosis. ○ Peak of curve is near 100 fL = majority of RBC in this specimen If MCV is normal= NO ANISOCYTOSIS is 100 fl in volume If MCV is low or high= WITH ANISOCYTOSIS ○ Normocytic = no anisocytosis, but has variation in size Generally: ○ Low MCV = Microcytic ○ Normal MCV = Normocytic ○ High MCV = Macrocytic 3 USING THE RBC HISTOGRAM Blood cell histogram is a visual display of cell size (X-axis) and cell MCV: 96.8 fL, RDW (CV): 14.1% frequency or the number of cells (Y-axis) Note: The definitive indicator of anisocytosis is RDW ○ X-axis is represented in femtoliters (fL) Provided by many high-volume instruments to provide size b Microcytic RBC population distribution of the different cell populations Type of curve: SHIFT TO THE LEFT Automated hematology analyzers produce histograms for RBCs, INTERPRETATION: WBCs, and Platelets. ○ Peak of curve is near 50 fL Two parameters calculated from RBC histogram: MCV and RDW ○ Normal RDW = no anisocytosis Instruments being used count those cells with volume sizes between ○ MCV is low (< 80 fL) = Microcytic 36 fL and 360 fL as RBCs ○ Even if the RBCs are smaller than normal, and RDW is normal, RBC histogram can measure cells as small as 24 fL. (However, cells we can conclude that there is no anisocytosis. that are counted in the 24 to 36 fL range are not included in the RBC For as long as the RDW is normal, there is no count.) anisocytosis, regardless of the size. Leukocytes are present in the diluted fluid containing RBCs, but their numbers are statistically insignificant in the count. The instrument computer can be calibrated to compensate for the presence of leukocytes. If the leukocyte count is significantly elevated, the RBC histogram will be affected. MCV: 54.6 fL, RDW (CV): 13.2% There are instances wherein if there is anisocytosis, the MCV still An alien from Mars showed one straight line on the curve normalizes. with 100 fL The MCV on the histogram may slightly differ from the MCV ○ Interpretation: all of their RBCs are 100 fL and do not have measurement of the machine any size variation (which is not possible for humans) If the results are consistent, there are no problems. But if the ○ It is impossible for human RBCs to have the exact same size values of the measurement and curve are inconsistent, there so variation is normal ONLY IF within acceptable ranges might be problems with the specimen or the machine. (which is the spaces seen on histogram results) ○ Thus, anisocytosis = increased number of red cells with REMINDER! variation in size. Always remember the reference range ○ MCV = 80-100 fL c Macrocytic RBC population ○ RDW-CV = 11.5 - 14.5% (adults) Type of curve: SHIFT TO THE RIGHT INTERPRETATION: ○ Peak of the curve is on the right side or beyond 100 fL = majority of RBCs are larger than normal 11 of 26  ○ RDW is high (> 14.5%) = there is anisocytosis ○ RDW cannot be used to evaluate the RBC size, but it tells us ○ MCV is high (> 100 fL) = Macrocytic the presence of anisocytosis and how severe it is Most recent hematology analyzers have provided two methods to calculate the RDW - RDW-CV and RDW-SD RDW is not a measurement of size, it is merely a measurement of anisocytosis. RDW-CV (coefficient RDW-SD (standard MCV: 119.2 fL, RDW (CV): 23.9% of variation) deviation) Note: Both the width of the The actual measurement If the legs of the RBC histogram are: RBC distribution curve of the width of the RBC Based on ○ CLOSED = NO ANISOCYTOSIS (Figure A & B) and the mean RBC distribution curve in fL ○ OPENED (widened/flattened curve) = ANISOCYTOSIS size (femtoliters) (Figure C) Reference 11.5% to 14.5% 39 to 46 fL A wider or flattened curve on a histogram indicates Ranges (memorize) more variation in the size of the cells (The cell (adults) population is NOT homogeneous.) Earliest method Width of the curve is provided by the measured at the d Dimorphic RBC population hematology point that is 20% The graph displays 2 peaks, indicating a BIMODAL curve, which analyzers to above the baseline implies that there are two major populations of RBCs in the sample. measure red cell NOT influenced by Examples of cases that may cause a bimodal distribution curve variations the MCV include the following: Remarks Dependent on the Better and more a. Recent blood transfusion (when normocytic donor width of the reliable measure erythrocytes are transfused to a recipient with microcytic red distribution curve of erythrocyte cells) and the MCV variability, ○ RBC population 1 is the patient’s original RBCs while specifically in highly population 2 is from transfused blood abnormal b. Cold agglutinin disease conditions. c. Hemolytic anemia with schistocytes present INTERPRETATION: ○ 2 peaks = BIMODAL 1st peak: 50 fL, 2nd peak: 100 fL ○ RDW is high = there is anisocytosis ○ MCV is normal MCV: 80.2 fL, RDW (CV): 37.2% Reference range for newborns: 14.2% to 19.9% ○ RDW is markedly increased in newborns but gradually, the Does bimodal curve automatically indicate anisocytosis? value will decrease until it reaches adult levels by 6 months of Answer: YES, as per the definition of anisocytosis, increased number age. of variation in the size of RBCs If the RBC histogram is wider than normal, the RDW would be ABNORMAL. 4 USING THE RDW VALUE Red Cell Distribution Width ○ A calculated index (from the RBC histogram) given by hematology analyzers to help identify anisocytosis and provide information about its degree. 12 of 26  SUMMARY Evaluate RBC Detect anisocytosis Size NORMOCYTIC PBS ✓ ✓ (Normal MCV and RDW) MCV ✓ ✓ RBC Histogram ✓ ✓ RDW X ✓ PBS can be used to evaluate the size of RBC and detect MACROCYTIC anisocytosis (High MCV and MCV can be used to evaluate the RBC size and detect anisocytosis RDW) ○ Low MCV (100 fL) = Macrocytic ○ Usually, if the MCV is low, there is also anisocytosis. However, it is not always the case because the MCV can be normal, but anisocytosis may occur. RBC Histogram can be used to evaluate RBC size and detect anisocytosis MICROCYTIC ○ Shift to the left = Microcytic (Low MCV and ○ Shift to the right = Macrocytic High RDW) ○ Flattened or widened histogram = anisocytosis/ more variation in size RDW cannot be used to evaluate the RBC size, but it can be used to detect anisocytosis Notes: ○ Low RDW (14.5%) = with anisocytosis. Low RDW has no clinical significance. ○ Normal RDW = RBCs may be small or large, but no Increase RDW indicates presence of anisocytosis. anisocytosis Normal RDW may indicate that the RBCs may be small or large, 2 ANISOCHROMIA but has no anisocytosis General term for a variation in the normal coloration (Turgeon) RDW is not necessarily related to RBC size, so even if MVC are ○ Normal color of erythrocyte is salmon pink abnormal, the RDW could still appear normal. ○ Other definition: Variation of the color of erythrocytes caused by unequal hemoglobin concentration Examples of conditions with their MCV and RDW values Normal RBC central area of pallor: 1/3 the diameter RBC size (as defined by MCV) May also mean the occurrence of hypochromic cells and Red Cell Decreased normochromic cells in the same blood smear (Henry’s) Normal MCV Increased MCV Distribution MCV (normocytic) (macrocytic) ○ May be found in sideroblastic anemias, also in a hypochromic Width (RDW) (microcytic) anemia after transfusion with normal cells and some weeks Anemia of after iron therapy for iron deficiency anemia NORMAL (little or chronic G6PD Liver disease no anisocytosis) inflammation deficiency a Hypochromic cells (ACI) INCREASED Iron deficiency Sickle cell Megaloblastic Central pallor is more than 1/3 in diameter or (anisocytosis) anemia (IDA) anemia anemia pale overall appearance Usually microcytic ACI has ↓ MCV and normal RDW, meaning the RBCs are small but ○ In hypochromic cells, there is a decreased consistent in size. concentration of hemoglobin, leading to a In contrast, in IDA, there's a ↓ MCV ↑ RDW, indicates that the RBCs paler appearance (less color). are small but vary in size, indicating anisocytosis ○ In the case of hypochromia, inadequate iron stores result in a ○ Regardless of the size, as long as the RDW is normal there is decrease in hemoglobin synthesis. Since hemoglobin is a no presence of anisocytosis critical component of red blood cells, insufficient production of ○ Regardless of the size, as long as the RDW is increase there hemoglobin often results in smaller-than-normal cells, or is presence of anisocytosis microcytosis. 13 of 26  GRADING OF HYPOCHROMIA The immune disorders that have spherocytes also are usually 1+ Area of central pallor = ½ of diameter characterized by a positive (+) result on the DAT. 2+ Area of central pallor = ⅔ of diameter 3+ Area of central pallor = ¾ of diameter BOARD EXAM QUESTION! 4+ Thin rim of hemoglobin (anulocyte) What is the color of the anti-human globulin reagent? a. Blue (used as anti-A) ANULOCYTE b. Yellow (used as anti-B) Also known as Pessary Cell / Ghost Cell c. Green Refers to a RBC demonstrating a thin rim of hemoglobin and a d. None of the above large, clear center Answer: GREEN–the blue reagent is the anti-A antisera while the May be observed in iron deficiency anemia (IDA) yellow reagent is the anti-B antisera. Grading: 4+ Note that a single term may be used for more than one meaning–the TESTS FOR HEREDITARY SPHEROCYTOSIS term ”ghost cell”, in clinical microscopy, pertains to RBCs in AUTOHEMOLYSIS TEST: greatly increased hypotonic urine. ○ This can be corrected with either glucose or ATP. OSMOTIC FRAGILITY TEST (OFT): increased osmotic fragility b Hyperchromic cells ○ NOT diagnostic of HS RBCs that lack central pallor even though ○ Heparin is the anticoagulant used for this test. they lie in a desirable area for evaluation EMA BINDING TEST: decreased fluorescence ○ The normal area that a central pallor ○ Considered as a flow cytometry assay occupies is ⅓ of the cell. ○ EMA: Eosin-5’-maleimide These RBCs are actually caused by a shape ○ Confirmatory test for Hereditary Spherocytosis change such as that found in spherocytes. c Polychromatophilic Erythrocytes ○ Therefore, when your red cells are hyperchromic, they are most Other terms: likekly to be spherocytic as well. ○ Diffusely basophilic erythrocyte (Wright-stained) True hyperchromia occurs when MCHC is HIGH ○ Reticulocyte (supravital stain–BCB or NMB-stained) Used in tandem–spherocytic, hyperchromic Larger than normal red cells with bluish tinge in Wright’s stain ○ The bluish tinge is caused by the presence of residual RNA HEREDITARY SPHEROCYTOSIS Large numbers are associated with decreased RBC survival, Characterized by spherocytes and hyperchromia hemorrhage or erythroid hyperplastic marrow Basically the ONLY disease in which the MCHC is high (above the reference range) GRADING OF POLYCHROMASIA ○ Some high MCHC are just falsely increased when seen in Grade Percentage of Polychromatophilic RBCs laboratory settings Slight 1% 1+ 3% 2+ 5% THREE CLINICAL MANIFESTATIONS OF SYMPTOMATIC 3+ 10% HEREDITARY SPHEROCYTOSIS 4+ >11% Splenomegaly–enlargement of the spleen Anemia 3 POIKILOCYTOSIS Jaundice Increased number of red cells with variation in SHAPE LABORATORY FINDINGS ○ Note that the normal shape of RBCs is a biconcave disk. DAT (direct antiglobulin test): negative Deviation from the discoid shape of an erythrocyte represents a ○ Note that spherocytes are not specific to hereditary chemical or physical alteration of either the cellular membrane or the spherocytosis (HS) and can appear in other immunologic physical contents of the cell. diseases. ○ However, HS can be differentiated using DAT as other diseases may test positive for it. MCV: normal to low MCH: normal MCHC: slightly increased (TRUE INCREASE) 14 of 26  DEHYDRATED HEREDITARY STOMATOCYTOSIS Also known Hereditary Xerocytosis The most common form of stomatocytosis characterized by the presence of XEROCYTES ○ Xerocytes are the dehydrated form of stomatocyte and appears to have puddled at one end (half light, half-dark) Even though the xerocytes are categorized as stomatocytes, it is not commonly called as stomatocytes, it is frequently called as it is a Spherocyte (xerocytes) because it has a different appearance than the rest. Almost spherical in shape and lacks central pallor Associted with: c Acanthocyte ○ Hereditary spherocytosis Also known as THORN CELL or SPUR CELL Confirmed through EMA Assay RBCs with IRREGULARLY spiculated surface ○ Autoimmune hemolytic anemia ○ The spicules arises in the cytoplasm are uneven. ○ Burns ○ This irregularity differentiates it from burr cells. ○ ABO Hemolytic Disease of Newborn (HDN) Unlike echinocytes, acanthocytes have fewer spicules. ○ Following transfusion of stored blood Associated with: ○ Abetalipoproteinemia ○ Spur Cell Anemia ○ McLeod Syndrome ○ Pyruvate kinase deficiency ○ Hepatic hemangioma ○ Neon