PRELIMS POST-LAB Flashcards
[?]is recommended; not shaking.
Inversion
Specimens are transported in an [?] to ensure complete clot formation and reduce agitation.
upright position
Samples of patients with known [?] shown be pre-warmed before testing.
Cold Agglutinin Antibodies
Specimens for routine testing should be delivered to the laboratory within [?] of collection.
45 minutes to 1 hour
Requirements of a Quality Specimen
Patient properly (?)
Patient properly (?) for draw
Specimen collected in the correct (?) and labelled correctly
Correct (?) used
Specimen properly mixed by (?) if required
Specimens not (?)
Specimens requiring (?) collected in a timely manner
Timed specimen drawn at the (?)
identified
prepared
order
anticoagulants and preservatives
inversion
hemolyzed
patient fasting
correct time
REASONS for SPECIMEN REJECTION
The test order (?) and the (?) identification do not match.
The (?) is unlabelled, or the labelling, including patient ID number, is incorrect.
The specimen is (?).
The specimen was collected at the (?).
The specimen is collected at the (?).
The specimen was (?), and the test requires whole blood.
The specimen was contaminated with (?).
The specimen is (?).
requisition; tube
tube
hemolyzed
wrong time
wrong tube
clotted
intravenous fluid
lipemic
Basic Hematologic Methods of Examination
QUALITATIVE Analysis
QUANTITATIVE Analysis
establishing morphological characteristics of cells in the blood, and blood – forming organs; and the presence of foreign elements in cells.
QUALITATIVE Analysis
giving the number of blood cells; concentration of hemoglobin, hematocrit, so as erythrocytic and thrombocytic indices.
QUANTITATIVE Analysis
The oxygen – binding capacity of blood is directly proportional to the [?] rather than the red blood cell count.
hemoglobin concentration
is the most important screening test for diseases associated with anemia and for following the response of these diseases to treatment.
HEMOGLOBINOMETRY
Makes use of whole blood or plasma
Methodologies
Difficult to crystallize and accurately weigh hb
Indirect methods
Gasometric Method Device
Van Slyke Apparatus
blood = oxygen
(standard conditions of temperature and pressure)
Gasometric Method
Gasometric Method
Oxygen:
Hemoglobin:
Oxygen: 1.34 ml
Hemoglobin: 1 gram
Measures only active hemoglobin
Gasometric Method
iron (blood) +hb
measurement of the iron content reflects the amount of hemoglobin in the blood
CHEMICAL METHOD
Iron - hb (sulfuric acid +potassium persulfate)
WONG TEST
Proteins are precipitated by (?), and the iron in the proteinfree filtrates is made to form
tungstic acid
ferric thiocyanate
measured spectrophotometrically for the amount of iron and thus hemoglobin
ferric thiocyanate
WONG TEST
iron: (?) gram per (?) gram hemoglobin or per ml of blood
0.347
100
100
WONG TEST American standard
0.338 gram per 100 gram hb/100 ml blood
The iron content of whole blood is determined
ASSENDELFT TEST
Iron - hb ( acid or ashing) =✓ titrated (TiCl3) =✓ complexed (developer) =✓ measured photometrically
ASSENDELFT TEST
Hemoglobin concentration is determined by assessment of blood specific gravity.
GRAVIMETRIC METHOD (Copper Sulfate Method)
GRAVIMETRIC METHOD uses CuSO4 solution made to a specific gravity of
1.053 or 1.055
drop sinks = SG…
equals or exceeds that of the CuSO4 solution
drop rises = SG…
is less
GRAVIMETRIC METHOD
Drops of blood are made to fall into a series of (?) having SG from (?) at an interval of
40 CuSO4 solutions
1.035 to 1.075
0.001
16 solutions may be used (?) with an interval of (?)
1.015 to 1.075
0.004
If the SG is the same as that of the solution, it will remain (?) for (?) before it sinks.
stationary
10 – 15 secs
VALUES :
(?) for women (corresponds approximately to (?)
(?) for men (corresponding to (?) o
Normal range : (?)
1.053; 12.5 g/dL
1.055; 13.5 g/dL
1.048 – 1.066
This is used for mass screening.
GRAVIMETRIC METHOD
Based on the use of colour standards with which the red colour of the whole blood is matched.
DIRECT MATCHING METHODS
- utilizes a printed colour scale graded from 10 to 100%
Tallqvist Hemoglobin Scale
- this is matched with the colour of a drop of the patient’s blood on absorbent paper.
Tallqvist Hemoglobin Scale
- highly erroneous because of subjectivity
Tallqvist Hemoglobin Scale
- the percentages on the scale are not accurate
Tallqvist Hemoglobin Scale
- Blood is drawn by capillary attraction between two glass plates, one transparent and the other white and translucent.
Dare Hemoglobinometer
- The colour is matched with a rotating disk on tinted glass varying in thickness and red color intensity
Dare Hemoglobinometer
- Comparison of the transmission of light through a layer of hemolyzed blood (oxyhemoglobin) of constant depth with that of a standardized glass wedge with a transmission of 540 nm (green colour)
Spencer Hemoglobinometer
- The intensity of light is measured rather than the colour.
Spencer Hemoglobinometer
The red cells are laked in dilute HCl, converting hemoglobin into acid hematin (brownish yellow color).
ACID HEMATIN
The preparation id diluted with distilled water until the color of the solution matches that of the comparator block.
ACID HEMATIN
The concentration of Hb is read directly from the gram scale etched on the tube
ACID HEMATIN
produce more homogenous lipid and protein moiety than does the acid hematin method, producing a more accurate solution.
Fairly strong alkali solutions
(COHb, SHb, Hi) are converted to hematin solution.
Inactive hemoglobin components
NaOH: 5 ml of 0.1 N
Blood: 0.05 ml of blood
-heated in a boiling water bath for 4 - 5 mins
-cooled
-read against a standard
ALKALI HEMATIN METHOD
(?) of blood + (?) Copper-free – glass – distilled aqueous NH4OH solution in a stoppered cuvette measured with a green filter at (?) nm with (?) as blank
0.02 ml
5 ml
540
0.007 N NH4OH
is measured as oxyhb at 415 nm, the SORET band of maximal absorbance for oxyhemoglobin.
Plasma hb
(absorbance band in the 400-430 nm region of the spectrum – which is actually the region of peak absorbance of heme compounds)
SORET band
Plasma hb is measured as oxyhb at 415 nm, the SORET band (absorbance band in the 400-430 nm region of the spectrum – which is actually the region of peak absorbance of heme compounds) of maximal absorbance for oxyhemoglobin.
HARBOE Method : ALTERNATIVE METHOD
Uses benzidine derivatives in which hb catalyzes the rapid oxidation of benzidine by hydrogen peroxide
METHOD of NAUMANN
More sensitive but it is inaccurate because normal plasma contains hydrogen peroxide inhibitor, and benzidine is carcinogenic
METHOD of NAUMANN
Blood is diluted in a solution of K(FeCN)6 and KCN.
CYANMETHEMOGLOBIN (HiCN) Method
The K(FeCN)6 oxidizes hb to Hi and KCN provides cyanide ions to form HEMIGLOBINCYANIDE (HiCN), which has a broad absorption maximum at a wavelength of 540 nm.
CYANMETHEMOGLOBIN (HiCN) Method
DRABKIN’S REAGENT
K(FeCN)6 = 0.200 g
KCN = 0.050 g
KH2PO4 = 0.140 g
Non – ionic detergent = 1 or 0.5 ml
Dilute to 1L with distilled water
20 ul of whole blood + 5.0 ml of DRABKIN’s Reagent
Incubate for atleast 3 mins at room temperature
Absorbance read at 540 nm
DRABKIN’S METHOD
Corrected by adding 20 ul of patient’s plasma to Drabkins reagent to be used as blank
Lipemia
Corrected by centrifuging test mixture and determining the hb in the supernatant fluid
Extremely high WBC counts (>30 x 109/L)
These provide resistance of cells to hemolysis causing now turbidity.
Hb S and Hb C
Correction is done by diluting Hb mixture 1:2 by taking 1 part and adding one part of distilled water.
Hb S and Hb C
The absorbance reading must be multiplied by the dilution factor which is 2.
Hb S and Hb C
Corrected by replacing NaHCO3 with KH2PO4
Easily precipitated globulins (WM, MM)
(?) in heavy smokers prolongs formation of HiCN to 1 hour. Readings taken at the normal pace cause erroneous results
Carboxyhemoglobin
Denotes the percentage of erythrocytes in a known volume of whole blood centrifuged at a constant speed for a constant period of time.
Hematocrit
A small amount of whole blood is centrifuged to determine maximum packing of erythrocytes, expressed as PCV or Hct.
MICROHEMATOCRIT METHOD
MICROHEMATOCRIT METHOD SPECIMEN REQUIREMENTS
Anticoagulated Whole Blood (EDTA)
Capillary blood collected in Heparinized Capillary Tubes
Specimens should be stored at room temperature and be processed within 6 hours after blood collection.
MICROHEMATOCRIT METHOD REAGENTS and EQUIPMENTS
Capillary hematocrit tubes (with red band or plain)
Non-absorbent sealing clay
Microhematocrit reader device
Microfuge with an RCF of 10,000 to 15,000 g
PROCEDURE
1. Fill atleast two capillary tubes approximately (?) full.
2. (?) the unfilled end.
3. (?).
4. Determine microhematocrit value using the (?)
5. Results should agree within (?) for duplicates.
2/3
Seal
Centrifuge
microhematocrit reader
0.02L/L
MICROHEMATOCRIT METHOD NORMAL VALUES
VARIATION dependent on age and gender of the population
o Birth :
o Age 1 :
o Adult Female :
o Adult Male :
o Birth : 45%-60%
o Age 1 : 27%-44%
o Adult Female : 37%-47%
o Adult Male : 40%-55%
MICROHEMATOCRIT METHOD
Significant Correlations: Increased
Polycythemia
Hemoconcentration due to shock
Dehydration
MICROHEMATOCRIT METHOD
Significant Correlations: Decreased
Anemia
Physiologic hydremias of pregnancy
MACROHEMATOCRIT METHOD: Specimen
Anticoagulated whole blood
o EDTA
o Ammonium Potassium Oxalate
MACROHEMATOCRIT METHOD PROCEDURE
Fill the (?) to 0 mark using a disposable pipette. The column should be bubble free
Place tube in the rack and set timer for
After 60 minutes, read using the (?) on the right. (10 – 0)
wintrobe tube
60 minutes
scale
1 hematocrit point =
1 hematocrit point =
0.34 gram hb/100 ml of blood
107,000 red blood cells/cumm of blood
applicable only to normocytic, normochromic red cells
Used as a random check
Rule of Three
MACROHEMATOCRIT METHOD BASIC EQUATIONS
3 x RBC = Hb
3 x Hb = Hct (+/- 3%)
decreases Hct values because of shrinkage of rbcs
Excess anticoagulant
may increase of decrease Hct depending on which part of the specimen (plasma or cells) is principally drawn into the microhct tube
Insufficient mixing of blood prior to obtaining Hct sample
decreases Hct because of leakage of specimen; erythrocyte loss is greater than plasma loss
Improper sealing of capillary tube
increases Hct. Results should be read within 10 mins after centrifugation.
Inadequate centrifugation or allowing tubes to stand too long after the centrifugation
increases the Hct.
Including a large buffy coat in the reading
increases or decreases the reading
Improper use of the microhematocrit reader
may damage the blood sample.
Heat sealing of capillary tubes
may cause the Hct to be falsely increased by as much as 0.02 L/L.
Trapped plasma
is the small amount of plasma that remains in the rbc portion of the spun. (sickle cell anemia, hypochromic anemia, spherocytosis, macrocytosis, and thalassemia)
Trapped plasma
(spherocytes and sickle cells interfere with rbc packing)
Certain abnormal rbc shapes
Hct and Hb appear normal; therefore, unreliable for evaluation of anemia immediately following acute blood loss.
During the first few hours of acute blood loss
which decreases plasma volume Specimen collection errors may alter the results
Dehydration increased Hct because of fluid loss
introduction of interstitial fluid to the sample causes falsely decreased Hct.
Hemoconcentration increases Hct
blood: 1:251 dilution
NH4OH: 0.007 N
OXYHEMOGLOBIN METHOD
The water used in the preparation of ammonia solution must
be glass distilled because minute amounts of copper in distilled water or other diluents employed will allow HbO2 to be converted to Hi and lower the values
OXYHEMOGLOBIN METHOD
ensures mixing and oxygenation of hemoglobin
Shaking
