Monospecific anti-human globulin (AHG) enables sensitized red cells to cross-link so that agglutination is visible.
Enhancement media are sometimes used to further promote agglutination and reduce incubation time.
- Low ionic strength saline (LISS) is the most common enhancement media. LISS reduces the ionic strength in the testing sample and causes reduction of the zeta potential. It increases antibody uptake and decreases incubation time. Gel technology, which is widely used, operates in a LISS environment.
- Polyethylene glycol (PEG): brings red blood cells (RBCs) closer together and concentrates antibodies by removing water molecules from the testing sample. It is the most sensitive of the enhancement media; strengthening almost all clinically significant antibodies. However, it will also enhance some clinically insignificant antibodies as well. Centrifugation should be avoided when PEG is used. PEG can cause aggregates to form if the sample (red cell - serum mixture) with PEG added is centrifuged. Reaction readings should only be done at the AHG phase. PEG is very effective for enhancing the reactions of antibodies in the Kidd blood group system, which are typically weakly reactive.
- 22% albumin: reduces zeta potential, bringing the RBCs closer together and enhancing agglutination. Although not widely used for routine testing, it is still an effective enhancement medium, especially with antibodies in the Rh system.
Detection of some IgG antibodies can be enhanced with enzyme test methods.
Proteolytic enzymes (eg, papain and ficin): denature some RBC antigens and remove negative charges from the RBC membranes. This reduces the zeta potential, bringing the cells closer together, enhancing some blood group antibody-antigen reactions. Enzyme techniques are particularly useful in the identification of Rh antibodies and antibodies in the Kidd, Lewis, P, and I systems. However, enzymes reduce or destroy the reactivity of other antigens including Fya, Fyb, M, and N. The effect of proteolytic enzymes on the S and s antigens are variable.
Technically, enzymes are categorized as “chemicals” rather than potentiating agents in most blood bank references. A number of other “chemicals” are available commercially for use in antibody identification and other blood banking procedures including: dithiothreitol (DTT), EDTA/glycine/acid (EGA), and chloroquine diphosphate. Regardless of which reagents are chosen for blood bank serologic testing, it is important for the technologist to be aware of the advantages and disadvantages of the reagent. Manufacturers’ directions must be followed and suitable controls included as appropriate.