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Exam Questions And Answers
Explain the structure of a red bell cell hemoglobin. - correct answer ✅It is composed of a globin with 2 alpha and 2 beta chains. Each chain contains a Heme group which contains a central Fe in which four oxygen group can bind What is porphyira? - correct answer ✅A abnormal synthesis of heme What are four hemoglobin derivatives and describe details of them? - correct answer ✅1. Oxyhemoglobin; oxygen attaches to Fe2+ replacing the water molecule. In order to carry the O2, Fe is kept in ferrous state my methemoglobin-cytochrome C reductase
- Carboxyhemoglobin: CO has a 200x higher affinity to hemeglobin than O2.
- Methemoglobin: ferrous Fe becomes ferric (Fe3+). MetHb cannot bind to oxygen making people with MetHb cyanotic (blue/purple discoloration)
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- Sulfhemoglobin: triggered by increase in fulfur content from sulfur containing drugs or chronic constipation. Process cannot be reversed and must be removed How does MetHb form? How is it identified? - correct answer ✅Ingestion of nitrates induces rapid oxidation of Heme that methemoglobin reducatase cannot keep up. MetHb can be identified by its characteristic band between 630-633nm in Evelyn-Malloy method What are the characteristic colors of the hemoglobin derivatives? How can different hemoglobins be identified? - correct answer ✅1. HbO2 = bright red
- HbCO = cherry pink Hemoglobin derivative can be differentiated by spectrophotometry or gas chromoatography What are the three types of Hb variants? Why do these occur? - correct answer ✅1. Hb-A1 (composed of alpha and beta chains)
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How can Hb-C be differentiatied fro other Hbs? - correct answer ✅Hb-C's negative charge is replaced by a positive charge and will become cathodal on electrophoresis Tetragonal crystals can form in the RBC making them rigid How can Hb-F be differentiated from other Hbs? - correct answer ✅Alkali denaturation test. Combining the Hb solution with KOH lowers the pH and precipates denatured Hb. Hb-F resists denaturation by alkali What use the main tool used for abnomral hemoglobin identification? How does it work? What determines the speed? - correct answer ✅Hemoglobin electrophoresis at a ph of 8.4. It separates hemoglobins based on isoelectric point (just like IEF). The movement speed is proportional to the difference between the ph of the electrophoresis and the pI of the hemeglobin.
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Ex. Hb-A (pI = 6.87) will move faster in a ph 8.4 buffer than Hb-C (pI = 7.3) What are the primary methods of hemeglobin measurement and what do they measure? - correct answer ✅The most common methods of hemeglobin measurement are blood oxygen capacity and blood iron content. Blood oxygen capacity measure fucntional Hb only. The oxygen capacity of Hb is 1.34 mg/g Blood iron content considers total iron content of the blood as bound to the Hb. The blood iron content is 3.47 mg/dL What are the secondary methods for hemoglobin measurement? How does it work? What's the procedure? - correct answer ✅Cyanmethemoglobin method (HiCN) is an estimation of all forms of Hb except for Hb-S. Drabkin solution is a combination of potassium ferricyanide and NaCN. The ferricyanide converts the Hb to MetHB. KCN (single rearrangement from NaCN to KCN) converts MetHb to
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b. Circulating haptoglobin can combine with approx 3g of hemoglobin c. In hemolytic anemias, haptoglobin-hemoglobin complexes are rapidly removed resulting in a decrease in serum haptoglobin d. Haptoglobin (as an acute phase reactant) is increased in inflammation when plasma hemoglobin breaks down, indirect- reacting bilirubin increases How can haptoglobin be seen when used as an acute phase reactant? - correct answer ✅In inflammatory events, haptoglobuin is increased when plasma hemoglobin breaks down and indirect reacting bilirubin increases This can been seen as a heme fraction, bound to albumin and methmalbumin and appears in the plasma using spectroscopic detection 9Schumm test) or my adding yellow ammonium sulfide to plasma containing methmalbumin which produces hemochromagen Hemoglobin also is broken down in kidney tubules, hemosiderin apears in urine. It appears as yellow-brown grandules either free
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flowing or within tubular epithelial cells/casts. Using Perl's stain, they become blue granules with Prussian blue What are tests for plasma Hb? - correct answer ✅Incubate benzidine compound and hydrogen peroxide and let stand for 10 min
- Hb acts as a catalyst for hydrogen peroxide to oxidize armoatic diamine -OD should be 0.1-0.
- for high levels of plasma Hb mearusre by whole blood cyanmethemogolbin method What is myoglobin? - correct answer ✅Its a conjugated protein resembling hemoglobin composed of 1 polypeptide chain and 1 heme which totals in 16000 mol weight (1/4 mol weight of Hb) How to separate Mb from Hb? - correct answer ✅1. Place sample in 80% ammonium sulfate. Solublizes Mb but not Hb
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What do the different layers mean in the hematocrit? - correct answer ✅1. Top of the tube = fatty layer
- normally is barely visible but becoms obvious in the presence of lipidemia
- Second layer = plasma -normally yellow and clear
- excessive hemolysis = cherry red color
- jaundice = yellow color
- Third layer = buffer coat
- normally less than 1mm thick
- if greater than 2%, recorded it as packed WBCs
- packed platlets are also found here
- Bottom layer = packed red cells -hematocrit reads this packed layer For microhematocrit, why are heparinized tubes used for fingertip blood and not for anticogulant venous blood? -
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correct answer ✅Fingertip blood can contaminate the sample decreasing the hematocrit by increasing coagulation. Heparinized tubes introduce an anticoagluant How do you perform a microhematocrit? - correct answer ✅1. Use heparinized tubes if acquiring fingertip blood otherwise use plain tubes
- Centrifuge at 14,490g
- Read the PCV with a microhematocrit reading device within 10 mins or the RBC layer will slant leading to a false result
- Duplicate determination must be recorded What is the erythocyte sedmentation rate? What does it depend on? - correct answer ✅ESR is the rate at which RBCs fall through the plasma producing a sediment depends on plasma fibrinogen concentration and the concentration of certain globulins What are the average ESR rates in men and women and when is the ESR normal/abnormal? - correct answer ✅ESR (men) 0-15mm/hr
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What other factors can affect ESR (not necessarily lead in the decrease?) - correct answer ✅1.Hypofibringenemia
- Polycythemia (increased Hb concentration)
- Abnormal Hb shape (sickle cell, spherocytes (smaller and denser), bizarre poikilocytes) Age, sex, and drugs (corticosteroids and contraceptives) How are RBC counts obtained with a hemacytometer? What caluclations are needed? - correct answer ✅RBC sample is loaded into teh RBC diluting pipet (at 1;200) and applied to a slide and placed on Using the center square of the hemacytometer count the RBC in the four corners and then the center of the square red cells/uL = number of cells counted x 200 (dilution correction) divided by 10 (depth) x 5 (area correction)
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What are the principles employed for automated particle counting?
correct answer ✅1. Electrical impedance: suspended in electrolyte solution passes through an electrical zone producing a pulse proportional to the volume of the cell
- Radio frequency: high frequency current penetrates the internal structure of the cells which is proportional to the nuclear size and density of the cell
- Light scatter: a pattern of light scatter is generated as it moves through a laser beam. This is proportional to its granularity and its reflectivity How do scatter plots use the principles for automated particle counting? - correct answer ✅They use discriminant function (DF) views 1, 2, or 3 DF1 = volume (y axis), scatter/light (x axis) DF2 = volume (y axis), conductivity/radio frequency (x axis) DF3 = volume (y axis), opacity ( x axis)
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List the different hematological stains and what they look for? - correct answer ✅1. Prussian Blue - stains for Iron ----potassium ferrocyanide + HCl ----site of non heme Fe appears deep blue green, cell nuclei is red
- Sudan Black B (SBB) - stains for phospholipids and sterols ----appears to stain both azurophilic granules and specific granules in neutrophils
- Crystal violet/methylene blue - stains for Heinz bodies (not viewable with Wright stains)
- Naphthol AS-D chloracetate (D-Cl-E) - stains for specific esterases ----specific for granulocytes and precursors that are positive ----differentiates AML from ALL
- Alpha-naphthyl acetate/alpha-naphthyl butyrate with NaF inhibitor ----NaF inhibitor reactions in monocytes, megakaryotes, platelets, and plasma cells but not granulocytes decrease from 3+ to 0-1+
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- Periodic acid-Schiff (PAS) - stains for glycogen
- Oil red O (ORO) - stain for neutral lipids
- LAP - stains for leukocyte alkaline phosphatase of neutrophils
- Acid phosphatase (ACP) - red granules in cytoplasm indicate acid phosphatase activity Summary PAS and ORO - used in acute lymphocytic leukemia (ALL) SBB, MP, D-Cl-E: used to rule in granulocytic leukemia Neutrophilic Alkaline Phosphatase (NAP): used to distinguish chronic myeloid leukemia (CML) from other leukemia ACP: rule in hairy cell leukemia or T cell ALL Explain the process of marrow erythropoiesis - correct answer ✅With RBC matruation, the size of the cell decreases and well as the disappearance of the nucleus
- Rubriblast
- Prorubricyte
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- Neurological symptoms (peripheral paresthesias/slow circulation in limbs)
- Previous medications (trasfusions/B12 injections)
- Underlying diseases (leukemia, etc)
- Physical findings ---skin pallor (pallor = non-hematological anemia, jaundice = hemolysis) ---eyes (sausage link veins) ---mouth (lead lines on gums, smooth tongue (glossitis) ---lymph nodes (infectious disease, leukemia) ---heart (murmurs, enlargement) ---spleen (mild to massive enlargement ---liver (alcoholism) ---bone (sternal pain = leukemia, back pain = myeloma) What is a "biscuit" cell? What disease is associted with it? - correct answer ✅A folded RBC. Diseases: Hb-C, Hb-SC
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What is a bronze elliptocyte and what disease is it associated with?
correct answer ✅A bipolar (on the ends) or central distribution of Hb Found commonly in sickle cell anemia What is a burr cell (echinocyte) and what disease is associated with it? - correct answer ✅The presence of small cells or fragments bearing one or a few spines Can occur in: --renal transplants --uremia --microangiopathic anemia --bleeding peptic ulcer --carcinoma
