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Total volume of blood in males 5-6L
Total volume of blood in females 4-5L
3 layers of buffy coat Uppermost: platelets / middle: Agranulocyte / lower layer: Granulocyte, nRBC
Blood pH 7.35-7.45 (ave. 7.40)
Whole blood spe. grav: 1.045-1.066
Serum spe. Grav: 1.024-1.028
Plasma SG: 1.025-1.029
Concentration of solutes dissolved in the blood Osmolality
Ref range for osmolality: 281-297 milliosmoles/kg
Blood makes ____ per body weight. 7-8% (75-85mL)
Most important function of blood: Respiratory function
Optimum conc. of EDTA anticoagulant in blood: 1.2-1.5 mg/dL
EDTA in dry form: Na²EDTA / K²EDTA
EDTA in liquid form: K³ EDTA
Anticoagulant choice in Hematology: EDTA
Disodium salt also known as ______. Versene 2
Tripotassium salt also known as _____. Sequestrene 3
EDTA actions to chelate ______. Calcium
_____ are recommended by the International Council for Standardization in Hematology (ISCH) abd CLSI as the anticoag. of choice for BLOOD CELL COUNTING AND SIZING because they produce less shrinkage of RBCs and less of an increase in cell vol in standing. K² EDTA
If conc. of EDTA exceeds 2mg/dL of WB, platelets may ______. Swell and Fragment (causing false higher account)
Platelet Satellitosis is caused by _____. Using EDTA as anticoag. causing false decrease PLT count.
Platelet satellitosis can be fixed by _____. Using sodium citrate
Increase EDTA is caused by _______. Underfilled / Short draw ( falsely low HCT/ESR)
EDTA cannot be used for coag studies because _____. Factor V is not stable / inhibits fibrinogen - thrombin reaction
Sodium Citrate action is to ______. Bind calcium to form soluble complex.
Anticoagulant of choice for coag studies: Sodium citrate
Ratio of blood to anticoagulant is (Na citrate) _____. 9:1
Sodium citrate should be buffered and use in concentration of ______. 3.2% or 0.109 M
Sodium citrate preserve factors ______. V and VIII
Anticoagulant for Standard Westergren ESR (black top): Sodium citrate
Citrate to blood ratio for Standard Westergren: 4:1
Sodium citrate conc. for standard Westergren: 3.8 %
Underfilled/short draw of citrated blood can cause _____. Prolong PT and PTT
In polycythemic patient there is hct of >55% which causes excess in citrate thereby prolonging coag test results. What is the remedy? Reduced the volume of citrate
Oxalate concentration: 1-2mg/dL
Oxalate actions to _____. form insoluble calcium oxalate
Forms of oxalate: Lithium oxalate / sodium oxalate / potassium oxalate / double oxalate
Oxalate form which is widely used : Potassium oxalate
Double balance oxalate : Potassium oxalate ( Paul Heller) causes cell shrinkage / ammonium oxlate (Wintrobe) causes cell swelling
Considered as a natural anticoagulant : Heparin
Anticoagulant of choice for most chemistry test Heparin (Lithium Heparin)
Optimal conc. of heparin 15-20 U/mL or 15-30 U/mL
Anticoagulant used for OFT and LAP test: Heparin
Heparin is not used for blood film preparation because: It produces a bluish background on Romanowsky stained smear
Heparin is not used for coag studies because It inhibits all stages of coagulation cascade specially thrombin
Anticoagulant for cardiopulmonary bypass patients Heparin
Used for preserving glucose / inhibitor of glycolysis / can also be used in determination of lactic acid and blood alcohol Sodium fluoride
Venipuncture Order of Draw: ETS or Syringe YELLOW (SPS) blood culture / BLUE (Sodium Citrate) / RED (serum tubes) with or without activators / GREEN (Heparin) / LAVENDER (EDTA) / WHITE (K² EDTA) / GRAY (Sodium Fluoride)
Skin Puncture Order of Draw Blood gases / SLIDES (unless from EDTA) / EDTA / other anticoagulated microtubes / non aticoagulated microtubes
Yellow stopper tubes could be: SPS (BLOOD CULTURES) first in order / or ACD (last in draw) for blood bank or in lympocytotoxicity testing
Depth of skin puncture: Not more than 2mm
Sites for skin puncture: Lateral (outside) or medial (inside) plantar (bottom) surface of the heel or big toe (INFANTS UNDER 1Y) / distal portion of the third or fourth finger on non dominant hand / earlobes (less pain, fewer nerve endings, less tissue juices)
In skin puncture, the phlebotomist should warm the site to a temp. no greater than 42°C for no longer than 3-5minutes
False increase in skin puncture: WBC count
False decrease in skin puncture: RBC count, Hematocrit, Hemoglobin and platelet counts
Most widely used for collecting venous sample ETS
Sterilization for ETS Gamma irradiation
Shelf life for ETS Atleast 1 year
Most preferred veins for venipuncture Median cubital
Least preferred veins for venipuncture Basilic vein
Color of 18g Pink
Color of 21g Green
22g color Black
23g color Blue (children)
Standard needle for routine veni accdg to rodaks 21g
Standard needle for routine veni accdg to steinenger 20g
Length of needle 1-1.5inches
Venipuncture angle 15° or 15-30°
Tourniquet application 7.5 - 10cm (3-4inches) above veni site for 1min
Blood pressure cuff as tourniquet 60mmHg (40-60mmHg)
Most convenient and commonly used method for blood film preparation: Wedge Smear (25/ 30-40°/ 30-45°<)
Thick blood smear is used for Malarial detection
Distance from the edge of slide blood film 0.25mm or 1 inch to 0.5 inches
Thin smear increase pressure / decrease angle, size of blood, speed
Thick smear decrease pressure / increase angle, size of blood, speed
Size of drop of blood (blood film) 2-3mm
In polycythemic patients (increased hct), angle of the spreader must be lower to 25°
In anemic patients (decreased hct), angle of the spreader must be higher
In cross sectional counting of cells, blood film is moved to Side to side (H to T)
In longitudinal counting of cells, blood film is moved from Tail toward the head (T to H)
In battlement/track pattern/back and forth or serpentine counting of cells, blood film is moved to three consecutive horizontal edge field
length of film slide 2/3 to 3/4
Ideal shape for blood film Finger shape
Feather edge of blood film should have Rainbow appearance
A non supravital stain, a polychrome stain that is defined as any stain containing methylene blue or it's products of oxidation and a halogenated fluorescein dye: Romanowsky stain
Fixative used in blood films Methanol
Stain used in blood film Wright's stain (Methylene blue + Eosin Y)
Buffer used in blood film Aged distilled water or 0.05M sodium phosphate (pH 6.4-6.8)
Blood films should be stained after 2-3 hours after specimen collection
Stain used for malarial detection Giemsa stain
Supravital stain used for reticulocytes New Methylene Blue
Supravital stain used for demonstration of Heinz bodies and Hgb H New Methylene Blue
pH of stain used in malarial blood film: 7.2
pH of stain used in bone marrow blood stain: 6.8
A continuous regulated processof blood cell production that includes cell renewal, proliferation, differentiation, and maturation. Hematopoiesis
These cells are present in the first few hours after an ovum is fertilized / can develop into any human cell type, including development from embryo into fetus. Totipotential Stem Cell
Cells present in several days after fertilization / they cannot develop into a fetus Pluripotential Stem Cell
Derived from pluripotent stem cells / they are limited to specific types of cells to form tissues (ex bone marrow) Multipotential Stem Cell
Suggests that all blood cells are derived from a single progenitor stem cell called a pluripotent hematopoietic stem cell / most widely accepted theory Monophyletic theory
Suggests that each of the blood cell lineages is derived from its own unique stem cell Polyphyletic theory
An ideal environment of HSC is the allowance for: Self renewal / Proliferation / Differentiation / Apoptosis
Provide protective and nourishing environment to HSCs Stromal Cells
Hematopoietic Development: begins around 19th Day of embryonic development / formation of primitive erythroblast / embryonic hemoglobin Mesoblastic Stage / yolk sac
Phase of hematopoiesis which begins at 5th to 7th week of gestation / production of megakaryocyte / Hgb F is the predominant Hgb Hepatic Phase
Prior to the fifth month of fetal development, hematopoiesis begins at the bone marrow Medullary (Myeloid Phase)
Embryonic Hemoglobins: Gower I, Gower II, Portland
Gower I (Globin Chain Combination) 2 Epsilon + 2 Zeta
Gower II (Globin Chain Combination) 2 Alpha + 2 Epsilon
Portland (Globin Chain Combination) 2 Zeta + 2 Gamma
Fetal Hemoglobin (Globin Chain Combination) 2 Alpha + 2 Gamma
Adult Hemoglobin Hb A/A¹ (Globin Chain Combination) 2 Alpha + 2 Beta
Then adult red marrow is found only in: RSVP 🔥 (Ribs, Sternum (Shoulder Blade, Skull, (children), Vertebrae, Pelvic Bone (Proximal end of long bones)
Principal source of production in adult Sternum (other flat bones)
Hematopoietically active marrow consisting of the developing blood cells and their progenitors Red/Active marrow
Hematopoietically inactive marrow composed primarily of adipocytes (fat cells) with undifferentiated mesenchymal cells Yellow/Inactive Marrow
80-90% active marrow in Newborn
50-60% active marrow in Adult
The process of replacing the active marrow by adipocytes (yellow marrow) during development is called Retrogression
Normal myeloid to erythroid ratio: 2:1 , 3:1, 4:1
Excluded from the ME ratio Monocytes and Lymphocytes
Normocellular marrow has 30 to 70% HSCs
Tissue basophils Mast cells
Bone forming cells (waterbug or comet appearance) Osteoclasts
Bome resorption or destroying cells Osteoclasts
Most commonly used site for bone marrow smear collection Posterior iliac crest
Ocassional used site for bone marrow smear collection Sternum
Site for bone marrow smear for newborns and infants Upper end of tibial bone
Trephine (Core) Biopsy needles: Trephine biopsy needle / Jamshidi needle / Westerman-Jensen needle
Bone Marrow Aspirate needles: Aspiration needle / University of Illinois Sternal needle
Extramedullary hematopoiesis occurs in: Liver and Spleen
A procedure which requires counting atleast 500, and preferably 1000cells be counted for a marrow differential Bone Marrow Differential
Biggest cell in bone marrow: Megakaryocyte (0.1-3.0%)
Most numerous cells in bone marrow: Metamyelocyte / Juvenile
Two most common erythrocytic maturation stages: Polychromatophilic and Orthochromatic normoblast (fried egg appearance)
Largest lymphoid organ Spleen
Spleen stores ____ % of platelets. 30%
Spleen: contains lymphocytes, macrophage, and dendritic cells (antigen dependent lymphocytosis) White pulp
Cord of billroth, contains specialized macrophages for removal of the senescent RBCs Red pulp
Removal of mature RBC through phagocytosis that leads to eventual degradation of cell organelles / removal of aged/senescent RBCs Culling
Removal of cell inclusions Pitting
Hypersplenism leads to ________. Pancytopenia (false decrease)
Necrosis of spleen due to entrapment of sickled RBCs such as in case of sickle cell anemia Autosplenectomy
The absence of of splenic function (due to either surgical removal or radiation exposure) Asplenia
Splenectomy or Asplenia causes ______. Pancytosis (false increase)
Hematopoeitic stem cell marker (lymphoid and myeloid precursor) CD 34
Pre CALLA marker CD 10
Most reliable way to differentiate immature from a mature cell Chromatin pattern
Erythropoietin is manufactured in _____. Liver and Kidney
Erythropoietin is mainly produced in Peritubular cells of the kidney
First morphologically identifiable RBC precursor Pronormoblast
Erythropoietin major stimulant: Hypoxia
NC ratio is 8:1 / earliest recognizable precursor Pronormoblast / Rubriblast
Intense basophilic cytoplasm / NC ratio is 6:1 / nucleoli are usually not visible Basophilic normoblast / Prorubricyte
Last stage capable of mitosis / first stage of hemoglobin synthesis / 3rd precursor stage Polychromatophilic normoblast / rubricyte
Pyknotic nucleus (homogeneous blue black mass) / LAST NUCLEATED STAGE Orthochromic normoblast / metarubricyte
Enveloped extruded nucleus found in metarubricyte Pyrenocyte
Last stage capable of hemoglobin production / fine basophilic reticulum of RNA Reticulocyte/ Polychromatic erythrocyte or macrocyte/diffusely basophilic erythrocyte
Are recognizable on wrightstained blood smears by their larger size and increased blue tint / found on reticulocyte stage Stress reticulocytes
6-8 um / biconcave disc / 120 +- 20 days Mature Erythrocytes / Discocytes
Rubricyte nucleus pattern Checkered
Rubricyte cytoplasm pattern Muddy gray
Lymphocyte nucleus pattern Crushed velvet
Lymphocyte cytoplasm pattern Robin's blue
Up to ____ erythrocyte produced from a single rubriblast 16
_____ erythrocytes are produced from a single rubriblast (rodaks) 8-32
Vit B12 or folic deficiency is seen in _____. Pernicious anemia (megaloblastic)
Nucleus lags behind cytoplasm in maturation Megaloblastic anemia
Cytoplasm lags behind nucleus in maturation IDA
Oval macrocytes are seen in Pernicious anemia
Microcytic, hypochromic RBC are seen in IDA
Macrophages provides iron directly to the normoblast for hemoglobin synthesis Suckling pig phenomenon
RBCs are biconcave and average ____ in volume 90fl
Average surface area of RBC 140um
A normal RBC has increase deformability TRUE
Peripheral/Cytoskeletal protein responsible for increase deformability Spectrin
Decrease in spectrin leads to having spherocytes/bronze cells
Enzyme found in the lipid portion of RBCs: LCAT
90% anaerobic glycolysis / major energy provider Embden Meyerhof Pathway
Glucose enters the RBC without energy expenditure via the transmembrane protein _____. GLUT-1
Most common enzyme deficiency in Embden Meyerholf Pathway Pyruvate Kinase
10% Glycolysis (anaerobic) / provides reduced glutathione (GSH) to prevent oxidative denaturation of hemoglobin Hexose monophoshate Shunt/ Pentose Phosphate pathway
Most common inherited enzyme deficiency and associated with Heinz bodies in Hexose monophosphate pathway G6PD
Converts back methemoglobin back to normal hemoglobin using methemoglobin reductase enzyme (corrective mechanism) / cytochrome B5 reductase / methemogbin reductase pathway Methemoglobin reductase pathway
Involved for reducing ferric iron into ferrous state Cytochrome B5 reductase / methemoglobin reductase
Generates 2,3 DPG that decreases hemoglobin affinity to oxygen Rapoport Leubering Pathway
ODC has ____ shape. Sigmoid
Arterial blood 95% has ______ mmHg. 95
Venous blood 70% has _____ mmHg. 40
p50: 26.6mmHg Demonstration of the relationship between blood pH and oxygen affinity of Hb / presence of Hgb F (increase) and increase in myoglobin
Bohr Effect 90%
90% of RBC Destruction / also known as the macrophage mediated hemolysis Extravascular Hemolysis
____ mg of urobilinogen are excreted in urine/day 1-4mg
Converts uncojugated bili to conjugated: UDP-G
10% of RBC destruction / 3H's (Hemoglobinuria,Hemoglobinemia,Hemosiderinuria) Intravascular hemolysis
One gram of hemoglobin can carry ____ of oxygen. 1.34mL
One gram of hemoglobin can carry ____ of Iron. 3.47mg
Adult hemoglobin has _____ dimer of beta chains. 2
Hgb concentration in children: 12-15g/dL
Hgb concentration in adult male: 14-18g/dL
Hgb concentration in adult female: 12 to 15g/dL
Ferrous iron to pyrole rings ratio: 4:1
Alpha and Zeta Hgb is found on chromosome 16 (the rest are found on chromosome 11)
65% hemoglobin synthesis occurs in ______. immature nRBCs
35% hemoglobin synthesis occurs in ______. Reticulocytes
The most abundant transition metal in the body.
Normal daily diet contains _____ iron. 15mg
______ of iron is absorbed in the duodenum and upper jejunum. 1-2mg
Ferrous iron and taken up from the the intestinal lumen into the enterocytes by the iron transport protein _____. Divalent monotransporter 1 (DMT)
Iron maybe stored as ferritin in the enterocytes or exported into the circulation by another iron transport protein, ________. Ferroportin
Major iron transport protein Transferrin
Master regulatory hormone of systemic iron metabolism Hepcidin
Major storage form of iron Ferritin (soluble)
Minor storage form of iron Hemosiderin (insoluble)
Hb A normal reference range 95-97%
Hgb F ref range 1%
Hgb A² ref range 2%
Hemoglobin which has 200x more affinity for CO² Carboxyhemoglobin
Light sensitive and imparts a typical brilliant CHERRY RED COLOR Carboxyhemoglobin
Hemoglobin with ferric iron / chocolate brown discoloration Methemoglobin / Hemiglobin
Irreversible Hgb / green flurochrome / blood is mauve lavender / C. perfringens infection Sulfhemoglobin
Reference method for Hgb Cyanmethemoglobin / Hemiglobincyanide (HiCN) Method
Cyanmeth Hgb rgt: Modified Drabkin's rgt
Cyanmethemoglobin is measured at ______. 540nm
All hemoglobin can be measured except: Sulfhemoglobin
Turbidity of sample in Cyanmethemoglobin det.can cause: False elevated results
Dihydrogen potassium phosphate / allows the solution to be read after 3 mins
Original Drabkin's reagent contains _____. Sodium Bicarbonate (10-15mins)
Converts methemoglobin to cyanmethemoglobin Potassium cyanide
Converts Hgb (w/ferrous iron) to methemoglobin Potassium ferricyanide
Improves lysis of RBCs and decrease turbidity Non ionic detergent
Dilution used for cyanmethemoglobin det. using capillary whole blood: 1:251
How do you obtain a 1:251 dilution for cyanmethemoglobin det. using capillary blood: adding 0.2mL (20mL) of blood to 5mL of cyanmethemoglobin rgt
Let stand for __ at room temp to allow full conversion of Hgb to cyanmethHgb 10minutes
In standard hemoglobin curve, x axis (abscissa) correlates _____. Hgb value (g/dL
In Standard Hgb curve, Y axis or ordinate correlates _____. Optical density (absorbance)
CuSO4 spe. grav. should be _____. 1.053 to 1.055
In copper sulfate method, distance between drop of blood and solution is ___. 1 cm
In copper sulfate method, acceptable drop of blood will sink in solution within _____. 15 seconds if Hgb conc. is > or equal to 12.5 g/dL
Copper sulfate method is also known as Specific Gravity Method
Gasometric method is also known as Van Slyke Oxygen Capacity Method
Chemical method is also known as Kennedy's Wong's
Variation of cell size Anisocytosis
Assess anisocytosis RDW
Normal MCV 80-100fL
Normal RBC size 6-8um
RV for RDW 11.5% to 14.5%
Microcytic criteria RBC size: <6um / MCV: <80fL
Macrocytic criteria size: >8um / MCV: >100fL
variation in hemoglobin content Anisochromia
Normal RBCs have central pallor of approx. 1/3 in diameter Normochromic
MCHC NV 32-36%
Variation in rbc shape Poikilocytosis
Polychromasia or polychromatophilic cells are indicative of Immature/young RBCs / Severe Anemia
Seen in abetalipoproteinemia / McLeod Syndrome Acanthocytes / Spur Cells / Thorn Cells
Seen in having exposure to hypertonic solution / anemia associated with renal insuffieciency (uremia) Echinocyte / Crenated cells / Sea urchin cell / Burr cells
Seen in hemoglobinopathies, Thalassemia Target cell / Codocyte / Mexican Hat Cell
Seen in Rh null disease Stomatocytes / mouth cells
Seen in Hereditary elliptocytosis (Leach phenotype) / megaloblastic bone marrow Elliptocyte / Ovalocytes
Schistocytes with hornlike projections Keratocytes
Seen in DIC, microangiopathic hemolytic anemia, TTP Dacryocytes
RBCs fragment at 45°C (normally at 49°C) / severe burns Pyropoikilocytes
Seen in ptx having malaria Semilunar bodies/ half moon
Seen in Sickle cell anemia Drepanocytes / Sickle cell
Holly leaf formation SC disease
Pich cells / triangular shape with 2 central pallor associated in DIC / microangipathia Knizocytes
A thinner variant of codocyte Leptocyte
Remnants of DNA / positive in fuelgen reaction / seen in Megaloblastic anemia Howell Jolly bodies
Blue berry bagel appearance / dark blue granules / precipitation of ribosomes and RNA / seen in Lead Poisoning Basophilic stipplings / punctuate basophilia
Figure of eight / microtubules of mitotic spindle Cabot Rings
Precipitated or denatured Hb / a pitted golf ball / napthalene balls / moth balls / G6PD deficiency Heinz bodies
Resembles basophilic stipplings / unused iron deposits / stains with Perl's Prussian Blue / sideroblastic anemia Pappenheimer bodies / siderotic granules
Requires Brilliant Cresyl blue as a supravital stain Hgb H
Barr of Gold / Clam shell appearance Hb CC crystal
Crystalline fingerlike / quartzlike crystal / washington monument crystal Hb SC crystals
Nucleated RBC that contains non heme iron particles / excessive iron overload / sideroblastic anemia Ringed Sideroblast
Maltese cross appearance Babesia spp
causative agent of Nantucket fever Babesia microtii
Stack of coin pattern of RBC / seen in Multiple Myeloma / Waldenstrom macroglobulinemia / Hyperproteinemia Primary Atypical pneumonia is also known as
Walking pneumonia causative agent for Primary Atypical pneumonia
M. pneumoniae Clumping of RBC with no pattern / seen in Cold agglutinin disease / primary atypical pneumonia
Created by: oddreann
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