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HEMA 1: LEC > Hemoglobinometry- > Flashcards

Hemoglobinometry- Flashcards

1
Q

• Done separately or as part of routine CBC
• Relatively simple
• Performed quickly by the laboratory

A

Hemoglobin determination

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2
Q

Indications for Hb estimation

A

• Determine presence and severity of anemia
• Screening for polycythemia
• Response to specific therapy in anemia
• Estimation of red cell indices
• Selection of blood donors

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3
Q

WHO defines anemia as Hb levels of_____ in women and_____ in men

A

< 12 g/dL

<13 g/dL

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4
Q

Physiological Factors

A

•Age
•Gender
•Pregnancy
•Exercise
•Posture (from lying to sitting/standing)
•Altitude
•Smoking

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5
Q

Pathological Factors

A

• Testosterone deficiency
• Renal deficiency
• Myelodysplasia
• Diminished erythropoiesis
• BM suppression
• Blood loss

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6
Q

Normal Haemoglobin concentration in humans
• Men -
• Women(non pregnant) -

A

150-20 g/l OR 13-18 g/dL

135 +/- 15 g/l OR 11-16 g/dL

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7
Q

Pregnant women -
1st trimester
2nd trimester
3rd trimester

A

124-135 g/l

110-117 g/l

106-109 g/l

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8
Q

• Birth-______ OR _____

Lower up to______ by 2mos of age
and _______10 yrs of age

A

180 - 40 g/I OR 15-20 g/dL

10-14 g/dL

12-15 g/dL

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9
Q

Blood can be collected from 3 different sources:

A
  1. Capillary blood
  2. Venous blood
  3. Arterial blood
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10
Q

Methods of Hb Estimation

A

Visual Methods
Photoelectric Method
Gasometric Method

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11
Q

Visual Methods

A

• Specific Gravity
• Sahli’s Acid Hematin
• Tallquist Hemoglobin Color Chart
• WHO Hemoglobin Color Scale

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12
Q

Photoelectric Method

A

• Cyanmethemoglobin method

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13
Q

• Most accurate method for estimation of Hb.
• Recommended by _______

A

Cyanmethemoglobin Method

International Committee for Standardisation in Hematology (ICSH)

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14
Q

Why is cyanmethemoglobin method recommended

A
  • All forms of Hb are converted to cyanmethemoglobin (except sulthemoglobin)
  • Stable and reliable standard is available.
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15
Q

Cyanmethemoglobin Method - Equipment

A

• Photoelectric colorimeter or
spectrophotometer
• Sahlis pipette at 20 micro litre
• Pipette 5 ml

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16
Q

Hb Estimation by Cyanmethemoglobin Method

Reagents required
› pH of the solution-______
› Unstable if_____
› Stored at _____temperature in_____

A

DRABKIN’S SOLUTION

7.0-7.4

exposed to light

room temp

brown borosilicate bottles

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17
Q

Hb Estimation by Cyanmethemoglobin Method

Reagents required
› pH of the solution-______
› Unstable if_____
› Stored at _____temperature in_____

A

DRABKIN’S SOLUTION

7.0-7.4

exposed to light

room temp

brown borosilicate bottles

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18
Q

DRABKIN’S SOLUTION

› Solution should be_____ in color

A

clear and pale yellow in color

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19
Q

Hb Estimation by Cyanmethemoglobin Method

› Available commercially at a specific concentration
› Stored in a______ as it is photosensitive
› Exposure to light causes deterioration in the strength of the standard
› The concentration of the standard used in this demonstration is_____

A

Hb Standard solution

brown bottle

14.8 gm%

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20
Q
  1. The method starts with hemoglobin, which is in its ferrous (Fe²⁺) state in red blood cells.
  2. _______ is added to the sample, causing oxidation of hemoglobin. This oxidation converts hemoglobin (Fe²⁺) into_______, which has iron in the ferric (Fe³⁺) state.
  3. _______ is then added to the methemoglobin, forming_______. This compound is stable and has a distinct color.
  4. The concentration of cyanmethemoglobin in the sample is measured using a colorimeter or spectrophotometer at a specific wavelength (usually_____). The absorbance is directly proportional to the hemoglobin concentration in the sample.
A

K+ Ferricyanide; methemoglobin (Fe+3)

Potassium cyanide (K+ Cyanide); cyanmethemoglobin (HiCN)

540 nm

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21
Q

Cyanmethemoglobin

MECHANICAL SOURCES OF ERROR:

A

• Pipetting error.
• Use of dirty or scratched cuvettes.
• Use of deteriorated reagent.

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22
Q

Conditions
may affect Hemoglobin
Determination

A
  1. Specimen:
    -turbidity
    -hemolysis
    -lipemic
  2. Reagent precipitation due to abonormal proteins
  3. Hemoglobin type
23
Q

Cyanmethemoglobin Method
Advantages
• All forms of Hb except_______ are converted to hemiglobincyanide/cyanmethemoglobin (HiCN).
•_____ error is not there as no color matching is required.
• Cyanmethemoglobin solution is____ and it’s color does not fade with time so readings may not be taken immediately.
• Absorbance may be measured soon after dilution.
• A reliable and stable reference standard is available from World Health Organisation for direct comparison

A

sulphemoglobin

Visual

stable

24
Q

Cyanmethemoglobin Method
Disadvantages
• Diluted blood has to stand for a period of time to ensure complete conversion of Hb.
•______ is a poisonous substance and that is why Drabkin’s solution must never be pipetted by mouth.
• The rate of conversion of blood containing______ is slowed considerably. Prolonging the reaction time to_____ can overcome this problem.
• Abnormal plasma proteins cause turbidity when blood is diluted with
_______.
• A high leukocyte count also causes_____ on dilution of blood.
•______ the diluted blood can help overcome the turbidity.

A

Potassium cyanide

carboxyhemoglobin; 30min

Drabkin’s solution

turbidity

Centrifuging

25
Q

Gasometric Method
• Oxygen carrying capacity measured by_______

A

Van Slyke apparatus

26
Q

• Oxygen carrying capacity measured by Van Slyke apparatus

A

Gasometric Method

27
Q

Gasometric method

• Based on formula,____ of Hb carries____ of oxygen
It does not measure (3)

• Time-consuming and expensive.

• Result is____ percent less than other
methods.

A

1 gm; 1.34 ml

carboxyhemoglobin
sulfhemoglobin
methemoglobin

2

28
Q

Gasometric method

• Based on formula,____ of Hb carries____ of oxygen
It does not measure (3)

• Time-consuming and expensive.

• Result is____ percent less than other
methods.

A

1 gm; 1.34 ml

carboxyhemoglobin
sulfhemoglobin
methemoglobin

2

29
Q

• Rough estimate is made from specific gravity of blood

A

Specific Gravity method

30
Q

SG Method aka

A

• Copper sulfate technique.

31
Q

• Used in mass screening like selection of donors.

A

SG Method

32
Q

• Used in mass screening like selection of donors.

A

SG Method

33
Q

• Rapid and simple
• Commonly used in blood donor selection

A

Specific Gravity Method

34
Q

SG Method

• A drop of blood is allowed to fall in______ of specific gravity of_____ from a height of____

• Specific gravity is equivalent to______

• Drop gets covered with_____

• If drop sinks,specific gravity is higher than copper sulfate

A

copper sulphate solution; 1.053; 1 cm

12.5 grams/dl

copper proteinate

35
Q

Common Sources of Error in Specific Gravity Method

  1. hypergammaglobulinemia (e.g. multiple myeloma)
  2. air bubbles
  3. Use of inadequate height for dropping
A

= falsely high Hb level

= falsely low Hgb

= underestimating the Hgb concentration

36
Q

Common Sources of Error in Specific Gravity Method

A
  1. hypergammaglobulinemia (e.g. multiple myeloma)
  2. air bubbles
  3. Use of inadequate height for dropping
37
Q

Acid Hematin Method
APPARATUS
Sahli apparatus (haemoglobinometer
or haemometer) which is composed of :-

A

Sahli tube
Pippette
Colour comparator

38
Q

Acid hematin

Hemolysis of RBC releasing
hemoglobin which is then converted to

A

acid hematin (Ferriheme chloride)

39
Q

ACID HEMATIN

STEPS

Put _____ till the mark _____
Add_____ blood (using the micro tube of the pipette)
Shake for_____
HCl + Hb →_____(brown in color)
Add____ to the acid hematin drop by drop
Match colors at full arm length and against light

A

0.1 HCl; 10%

0.02 ml

15 minutes

acid hematin

distilled water

40
Q

ACID HEMATIN

Adding____ to the blood causes :-Hemolysis of RBCs

Formation of_____
[HCl + Hb → acid hematin (brown in color)]

The intensity of color is_____ to hemoglobin content in blood.

A

HCl

acid hematin

proportional

41
Q

Role of HCl

• As it is_____ results in the hemolysis of RBCs.

• Convert the hemoglobin to a brownish____

A

hypotonic

acid hematin

42
Q

PRECAUTIONS
1. Be sure that there is no_____ in the blood column.
2. Clean the tip of pippette from any blood to avoid _____results.
3. Withdraw the blood_____ from the finger
> Ensure alcohol is completely dried
Avoid compression of the finger

A

air bubbles

false high

horizontally

43
Q

Advantages of Acid Hematin Method

A

• Easy to perform
• Quick
• Inexpensive
• Can be used as a bedside procedure
• Does not require technical expertise

44
Q

Disadvantages of Acid Hematin Method

• For maximum color,______ is required
•_____ with brown glass standard is not possible
• (3) are not converted to acid hematin
• Development of color is slow and____ is not stable
•_____ will influence the comparison of colors

A

longer time

Perfect matching

Carboxyhemoglobin,methemoglobin and sulfhemoglobin

acid hematin

Source of light

45
Q

• Series of lithographed colors said to correspond to Hb values ranging from 10 to 100 percent

• Blood obtained from finger puncture

• Placed on a piece of absorbent paper

• Color is matched against the color on the chart

• Corresponding reading taken

• Cheap and simple

• Error ______percent

A

Tallquist Hemoglobin Chart

20 to 50

46
Q

WHO Hemoglobin Colour Scale

• Devised by

A

Scott and Lewis

47
Q

• Principle is similar to Tallquist method

A

WHO Hemoglobin Colour Scale

48
Q

WHO Hemoglobin Colour Scale

• Rapid,simple,inexpensive,reliable
• _______ for diagnosis of anemia
• Printed set of colors corresponding to Hb values from______
• Efficiency-greater than _____percent in detecting anemia
•____ percent-in classifying its grade

A

1 gram/dl

4-14 grams/dl

90

86

49
Q

Useful for screening blood donors
Screening women and children in health programmes
Iron-therapy

A

WHO Hemoglobin Colour Scale

50
Q

• Modification of cyanhemoglobin method
• Other chemicals:
-SLS (Sodium Lauryl Sulfate)
-imidazole
-SDS (Sodium Dodecyl Sulfate)

A

Automated Method

51
Q

• Measurements are made at various wavelengths depending on final stable product

A

Automated method

52
Q

• Absorbance measurements are made at a set time interval after mixing the blood and the active reagents, but before the reaction is complete.

A

Automated Method

53
Q

• The cyanide-free reagent is ideal for use on an automated hig
through-put clinical hematology analyzer.

A

Automated method

HEMA 1: LEC (20 decks)

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