Washing red blood cells and preparation of its suspension Compiled by: Thamer Hamdan M.Sc. Clinical Microbiology and Immunology

Objectives The student should be able to know the purposes of red blood cells washing. The student should perform the washing of red blood cells. The student should prepare red blood cells suspension.

3% Red Blood Cell Concentration in Saline Between 2-5% cell suspension provides optimum antigen concentration for the tube method for red blood cells typing. To make sure your suspension is within this range use reagent red cells for comparison. HOW TO OBTAIN THIS?

Washing Red Blood Cells before Making the 3% Suspension The purpose of washing the red blood cells is to remove plasma, microaggregates, cytokines, and unwanted antibodies that may interfere with antigen-antibody reaction.

The following may be in the plasma and may interfere with testing: Soluble antigens such as A and B may be present and neutralize your reagent. Interfering proteins such as Wharton's jelly that is seen in newborn cord blood, cold-acting autoimmune antibodies, and increased levels of immunoglobulins that may cause either agglutination or rouleaux. Hemolyzed red blood cells due to a difficult draw will interfere in your grading interpretation of hemolysis Fibrinogen can result in fibrin strands forming that makes grading reactions difficult.

Good Technique when washing and making a 3% cell suspension involves the following Place 1 to 3 drops of blood in the tube Aim the tip of the saline bottle towards the center of the tube and forcibly squirt saline into the tube. Fill the tube 3/4 full of saline (there will be less splattering in the centrifuge) Centrifuge long enough spin to pull most of cells into a button in the bottom of the tube. Decant the saline completely Shake the tube to resuspend cell button before washing the cells again. It will depend on the procedure being done as to how many washings are going to be done.

Two Methods of preparing Washed RBC: 1. Manual Washing - This uses 0.9% saline to wash RBCs by manual centrifugation. This method however is much more expensive than automated washing. 2. Automated Cell Processor - It uses 0.9% saline to wash the RBCs in a series of continues washes.

THAT`S ALL