10. Centrifuge the tube at 3000 rpm for 2–3 minutes.

11. After the centrifugation, remove the supernatant

without disturbing the red cell button.

12. Take 0.2/0.3 mL of packed red cells from the above

cell button and resuspend them in 10 mL of Erywell

solution for preparation of 2/3% red cell suspension.

To obtain a 5% red cell suspension resuspend 0.5 mL

of packed red cells from the above cell button in 10 mL

of Erywell solution.

13. Store the red cell suspension in a sterile 10 mL vial.

14. The red cell suspension so obtained is ready-to-use for

testing.

Storage

1. The whole blood preserved in Erywell solution

prepared by the quick method can be stored up to 4

weeks at 2–8°C.

2. The ready-to-use 2–3–5 % stabilized red blood cell

suspension in Erywell solution should be stored at

2–8°C and can be utilized for weeks from the date of

preparation.

Precautions

1. Store Erywell at 2–8°C with cap tightly closed.

2. Do not contaminate the solution as it may subsequently

affect the stability of red cell suspension.

3. Glassware used to retrieve Erywell red cell suspension

should be scrupulously clean and sterile.

ABO GROUPING

Slide ABO Grouping Test

1. Prepare a 10% suspension in saline of the red cells to

be tested.

2. Mark the left side of a clean glass slide ‘anti-A’ and the

right side ‘anti-B’.

3. Add to the left side a drop of anti-A grouping serum

and to the right side a drop of anti-B grouping serum.

4. Next to the antisera drops, place 1 drop of 10% saline

suspension of unknown red cells. The contents on the

left and right sides should not get mixed up.

5. With one-half of an applicator stick, mix the red cell

suspension with the anti-A serum; and with the other

half-mix the red cell suspension with the anti-B serum.

6. Gently rock the slide back and forth and observe the

mixture for one minute unless agglutination occurs

earlier.

7. It is necessary to confirm the slide grouping by

testing the unknown serum with the known red cell

suspensions of groups A, B, and O as indicated for

six-tube method. (Instead of 10% saline suspension,

citrated/oxalated blood or direct blood from skin

puncture may be taken. Slow and weak agglutinations

may occur in A subgroups).

Six-tube Method (Fig. 11.3)

1. Separate the serum from red cells or clot of unknown

specimen.

330 Concise Book of Medical Laboratory Technology: Methods and Interpretations 2. Prepare a 2% suspension in saline of the red cells of

the unknown specimen.

3. For each specimen, six test tubes are labeled showing

in addition to proper identification of the unknown

specimen the undermentioned information: Tube 1,

anti-A; tube 2, anti-B; tube 3, anti-AB; tube 4, A; tube

5, B; tube 6, O.

4. Place 1 drop of anti-A serum in tube 1, 1 drop of the

anti-B serum into tube 2, and 1 drop of the anti-A, B

serum into tube 3.

5. Add two drops each of the serum of the unknown

specimen to tubes 4, 5, and 6.

6. Add one drop each of the 2% red cell suspension in

saline of the unknown specimen to tubes 1, 2, and 3.

7. To tube 4, add 1 drop of a 2% red cell suspension in

saline of group A cells; to tube 5, add 1 drop of a 2%

red cell suspension in saline of group B cells; and to

tube 6, add 1 drop of a 2% suspension of red cells in

saline of group O cells.

8. Mix the contents of all tubes by shaking the test tube

rack.