Immunohematology - Red Cell Affinity Column Technology

by Nancy Westbrook, 2000

In preparing and updating Immunohematology lectures for the MLT students, I have become very excited about what is said to be the most pronounced change in blood bank testing methods in decades. This change involves the use of red cell affinity column technology or adherence technology. There are several similar systems on the market, but the one that I have been learning about is Gamma-React, an acronym for Red Cell Affinity Column Technology.

This system lends itself especially well to the performance of antibody screens, antibody panels, and crossmatches. ABO and Rh typing as well as phenotyping may also be done.

The principle of the testing procedure is based on the adherence of antibody coated red blood cells to an immunologically active material. Sensitized red blood cells are adsorbed to a ligand or binding substance, which is attached to an agarose gel. There are various ligands or proteins which can be attached to agarose, enabling antibodies to then be bound to its surface. Bacterial proteins that have been used as ligands include Protein A and Protein G which are derived from Staphlococcus aureus.

Two distinct layers of different types of gels are used in the system. This affords a clear separation of sensitized and non-sensitized red blood cells. In a positive reaction, the red blood cells are coated with IgG and bind to immunoreactive gel particles, which are antibodies to the IgG coating the cells. This occurs mostly at the top of the gel column. In a negative reaction, the red blood cells are not coated with antibody and pass through to the bottom of the gel column.

Equipment necessary for the system includes:

• A special type of centrifuge
• A special incubator
• A viewbox
• Microcolumn strips

Testing is performed in pre-filled microcolumns. These come in strips of 6 or 8. For example, in running an antibody screening test, 50 ul of serum or plasma and 50 ul of 0.8% cell suspension are added to the upper test chamber. The microcolumn strip is incubated at 37 degrees C for 15 minutes. The strip is then placed in a specifically designed centrifuge and spun through 3 cycles.

1. 15 seconds at 1000 g

The red blood cells are forced through the viscous barrier which separates the test chamber and the immunoreative agarose material.

2. 30 seconds at 400 g

The red blood cells are forced into the gel. If the red blood cells are coated with antibody, they will be bound to the anti-antibodies or immunoreactive particles.

3. 90 seconds at 1500 g

Any red blood cells, which did not become bound, are forced to the bottom. If some or all of the red blood cells remain at the top of the column, the results are positive. If all of the cells settle to the bottom, the results are negative.

The advantages of Red Cell Affinity Column Technology include:

1. Detection of IgG coated red blood cells. Fewer non-clinically significant antibodies are detected than with other similar techniques.
2. Reactions are sensitive and specific.
3. Both positive and negative reactions are clear.
4. Results are reproducible.
5. Results are stable for 24 hours.
6. Short centrifugation times are required.
7. Only a small workstation is needed.
8. No washing of cells is required.
9. There is no need for slides, tubes, saline for the cell washer, a cell washer, a microscope, an incubator, albumin, or antihuman globulin.

So this year, I am adding some very important information for Immunohematology students. Red cell Affinity Column Technology has the potential for making a tremendous difference in the way we do our testing in the Blood Bank in the future. In a way, it is sad to see the old procedures that have been with us for so long becoming obsolete, but actually, I think this procedure has too much potential to allow us to feel anything but very excited about it.

So if you have not already done so - give this technology a try.