LABORATORY ACTIVITY 11

Question 1

Normal hemoglobin function carries oxygen around the body so that the tissues could breathe. What happens is there is a production of abnormal hemoglobin called

Answer 1

hemoglobin S.

Question 2

● caused by amino acid substitution at the 6th position of the B chain

Answer 2

HEMOGLOBIN S (HBS)

Question 3

● Glutamate/Glutamine should be in the 6th position but it is replaced by valine

Answer 3

HEMOGLOBIN S (HBS)

Question 4

● RBC takes on sickle-like-shape when the oxygen supply is decreased

Answer 4

HEMOGLOBIN S (HBS)

Question 5

● Causes sickle cell anemia and sickle cell trait

Answer 5

HEMOGLOBIN S (HBS)

Question 6

There is lower HbS (?).

Answer 6

20-40%

Question 7

METHODS USED

Answer 7

Sodium Metabisulfite Method
Sodium Dithionite (Solubility) Test/Solubility Test

Question 8

● Deoxygenated hemoglobin

Answer 8

Sodium Metabisulfite Method

Question 9

● Uses saponin

Answer 9

Sodium Dithionite (Solubility) Test/Solubility Test

Question 10

○ Immediately lyse RBCs

Answer 10

saponin

Question 11

● Cannot differentiate between sickle cell anemia and trait

Answer 11

Sodium Metabisulfite Method

Question 12

● HbS and other sickling hemoglobins are positive

Answer 12

Sodium Dithionite (Solubility) Test/Solubility Test

Question 13

● Sickle cells/holly leaf form is considered positive
○ There is abnormal Hbs

Answer 13

Sodium Metabisulfite Method

Question 14

Problem: cannot differentiate sickle cell anemia and sickle cell trait. It cannot measure the degree of deoxygenation

Answer 14

Sodium Metabisulfite Method

Question 15

● Forms liquid crystals and turbid appearance is a positive result

Answer 15

Sodium Dithionite (Solubility) Test/Solubility Test

Question 16

PROCEDURES

Answer 16

A. SCRIVER AND WAUGH METHOD

B. SICKLE CELL SLIDE TEST

C. TUBE SOLUBILITY TEST

Question 17

A. SCRIVER AND WAUGH METHOD
1. Place a rubber band around the base of the middle finger and allow staying in place for [?]
2. Make a finger puncture on the ball of the finger and place a drop of [?] on a slide
3. Immediately cover with a coverslip and seal edges with [?]
4. Incubate the preparation at room temperature. Observe for red cell sickling at hourly intervals for [?] if desired.
5. Microscopic examination (400x): if more than [?] of the cells are sickled, the result is positive

Answer 17

5 minutes

capillary blood

petroleum jelly

2, 3 hours, or after 24 hours

10%

Question 18

● The finger may appear bluish/purplish and you may feel numb. This is due to [?] of the finger

Answer 18

deoxygenation

Question 19

● Because oxygen from the environment can [?] the blood

Answer 19

oxygenate

Question 20

● In the absence of the HbS solubility test, the [?] is useful in detecting sickle cells in patients who have either sickle cell disease or sickle cell trait.

Answer 20

sickle cell slide test

Question 21

1. Weigh [?] and transfer to a test tube capable of holding [?].
2. Add [?], stopper, and mix until the chemical is fully dissolved. The chemical can only be used within the day it was suspended (?)
3. Deliver one drop of patient’s [?] on a slide and add an equal volume of freshly made reagent, mix and cover with a cover glass. Exclude any air bubbles.
4. Place the slide in a plastic box or Petri dish with damp piece of [?] at the bottom to prevent drying of the preparation (moist chamber). Leave at room temperature.
5. After [?], examine the preparation microscopically for sickle cells. Focus the cells first with the [?] and examine for sickling using the [?]. Examine several fields. Sickling usually takes place in one part of the preparation rather than the other.
6. Sickle cells usually appear in [?]. Report as “sickle cell test positive” when crescent shape cells are seen, or ‘sickle cell test negative” when cells appear rounded or oval shape.

Answer 21

0.1 g of sodium metabisulphite; 15 ml of water

5 ml of distilled or deionized water; within 8 hrs

capillary blood or well mix venous blood

blotting paper or tissue

10-20 minutes; 10x objective; 40x objective

crescent shape with pointed ends or holly leaf shape

Question 22

The principle was based on turbidity created when Hb S is incubated with sodium dithionate.

Answer 22

solubility method

Question 23

C. TUBE SOLUBILITY TEST
1. Add [?] of blood with [?] of the reagent prepared in procedure B (?)
2. Mix and stand at room temperature for [?]
3. Examine the tubes using a [?] as the background in ambient light settings.
4. Interpret results as positive if the [?] are not visible.

Answer 23

20 uL; 2 ml; 2% w/v sodium metabisulphite or sodium dithionite

5 minutes

white board with black lines

black lines

Question 24

• sickling readily occurs when oxygen is reduced/slightly decreased

Answer 24

● In sickle cell anemia

Question 25

• Sickling ofRBCs (HbS): 80-90% (severe: causes stroke to the brain or death)

Answer 25

● In sickle cell anemia

Question 26

Some hemoglobins that aggregate and have (?) are capable of polymerizing and crystallizing within the red cell causing a distortion of cell shape (sickle shape).

Answer 26

reduced solubility

Question 27

Hb S (Sickling Hb), when fully
oxygenated is (?).

Answer 27

fully soluble

Question 28

Polymerization and formation into tactoid crystals occur only when (?) at tissue level.

Answer 28

oxygen is decreased

Question 29

Laboratory Activity 11: SICKLE CELL EXAMINATION

Specimen needed:

Answer 29

Capillary blood