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Practical Blood Bank

Practical Blood Bank. Adsorption. Lab 8. Adsorption. Antibody can be removed from a serum sample by adsorption to red cells carrying the corresponding antigen.

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Practical Blood Bank

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  1. Practical Blood Bank Adsorption Lab 8

  2. Adsorption • Antibody can be removed from a serum sample by adsorption to red cells carrying the corresponding antigen. • After the antibody attaches to the membrane-bound antigens and the serum and cells are separated, the specific antibody remains attached to the red cells. • It may be possible to harvest the bound antibody by elution.

  3. Adsorption techniques are useful in such situations as: • Separating multiple antibodies present in a single serum. • Removing autoantibody activity to permit detection of coexisting alloantibodies. • Removing unwanted antibody (often anti-A and/or anti-B) from serum that contains an antibody suitable for reagent use. • Confirming the presence of specific antigens on red cells through their ability to remove antibody of corresponding specificity from previously characterized serum. • Confirming the specificity of an antibody by showing that it can be adsorbed only to red cells of a particular blood group phenotype.

  4. Specimen • Serum or plasma containing antibody to be adsorbed. • Reagents • Red cells (eg, autologous or allogeneic) that carry the antigen corresponding to the antibody specificity to be adsorbed.

  5. Procedure • Wash the selected red cells at least three times with saline. • After the last wash, centrifuge the red cells at 800 to 1000 × g for at least 5 minutes and remove as much of the supernatant saline as possible. Additional saline may be removed by touching the red cell mass with a narrow piece of filter paper. • Mix appropriate volumes of the packed red cells and serum and incubate at the desired temperature for 30 to 60 minutes. • Mix the serum/cell mixture periodically throughout the incubation phase.

  6. Centrifuge the red cells at 800 to 1000 × g for 5 minutes to pack cells tightly. Centrifuge at the incubation temperature, if possible, to avoid dissociation of antibody from the red cell membranes. • Transfer the supernatant fluid, which is the adsorbed serum, to a clean test tube. If an eluate is to be prepared, save the red cells. • Test an aliquot of the adsorbed serum, preferably against an additional aliquot of the cells used for adsorption, to see if all antibody has been removed.

  7. Interpretation • If reactivity remains, the antibody has not been completely removed. No reactivity signifies that antibody has been completely adsorbed.

  8. Notes • Adsorption is more effective if the area of contact between the red cells and serum is large; use of a large bore test tube (13 mm or larger) is recommended. • Multiple adsorptions may be necessary to completely remove an antibody, but each successive adsorption increases the likelihood that the serum will be diluted and unadsorbed antibodies weakened. • Repeat adsorptions should use a fresh aliquot of cells and not the cells from the prior adsorption. • Enzyme pretreatment of adsorbing cells can be performed to increase antibody uptake for enzyme-resistant antigens.

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