Use of the EDTA-Glycine-Acid Method for Red Cell autoadsorption and phenotyping in patients with autoantibodies
A colleague in New Zealand reports that the New Zealand Blood Services is presently considering standardization of their approach to autoadsorption and red cell phenotyping of patients with autoantibodies. Currently, a majority of sites use the PEG method for autoadsorption and for red cell phenotyping. However, a few sites use the EDTA-Glycine-Acid method for red cell autoadsorption and/or phenotyping. The inquiring New Zealander wants to know if others have experience using EDTA-Glycine-Acid treated red cells for autoadsorption and phenotyping, and how reliable is phenotyping of these red cells, assuming the patient has not been recently transfused?
The following responses have been received.
ADDENDA Nov. 8, 2004
- A colleague in Massachusetts reports that their AABB-accredited reference lab uses the EDTA Glycine Acid (EGA) method to remove bound antibody in order to antigen type patients. Previously they used Chloroquine, but abandoned this because it did not remove all of the bound antibody. In their hands the EGA method works well at removing bound antibody from the red cells, is quick, easy and reliable. The only drawback is that Kell system antigens are destroyed. They do not use EGA for autoadsorption. Instead they treat the patient's cells with ZZAP.
- According to Marilyn Moulds, VP Education, Immucor-Gamma (attribution used with permission) "The EDTA-Glycine Acid method for removal of bound IgG from the red blood cells for phenotyping has been in use for quite a few years, and since a commercially prepared kit has become available, it is perhaps more widely used than before. It replaced Chloroquine diphosphate (except for when you have to type for Kell System Antigens, Bg and Era, which are destroyed or weakened) because it is more efficient at removing IgG and is faster. As always, there are limitations to every technique, but it does give reliable phenotyping results for the common red cell antigens. I think it is not widely used for preparing red cells for autoadsorption because of the small volume of cells that are retrieved in the procedure and other methods are more efficient (ZZAP, for instance, that also enzyme-treats the red cells in addition to removing bound IgG). Thus, I do not know of any data for the use of EGA-treated cells for autoadsorption, although I see no reason why it would not work. There is also some recent discussion on the use of PEG for autoadsorption, where some alloantibodies have been shown to be weakend by this procedure."
She adds that in her experience "PEG is NOT used in phenotyping" and she requests information from those who might be using that method for phenotyping. In summary, she states that two procedures are possibly needed, one for phenotyping red cells that have a positive DAT (EGA-treated and/or Chloroquine-treated cells) and one for autoadsorption [enzyme-treated, ZZAP-treated or WARM-treated, or PEG-adsorption (?) which has limitations]. She concludes saying "If one wants to do a study to see if EGA-treated cells can be effective in removing autoantibody, then we would have data to know if it is effective and economical."
- According to an experienced immunohematologist in Michigan, the use of PEG in adsorptions is problematic, since PEG precipitates immunoglobulins, which reduces reactivity of alloantibodies, if present. For this reason, his institution was unable to validate the PEG-adsorption procedure in his laboratory. (See the abstract from Annual AABB Meeting in Transfusion 2000;40(S):28). His laboratory uses a modified ZZAP procedure that utilizes ficin instead of cysteine-activated papain. For allogeneic adsorption in recently transfused patients, they attempt to ascertain patient's Rh and Jk phenotypes and adsorb with ZZAP-treated Rh and Jk phenotype identical RBCs. The method can be found in the AABB Technical Manual. At the 2004 AABB Meeting (Transfusion 2004;44(S):121), Graves and Pierce reported on their experience using ZZAP-treated RBCs and a 10-minute incubation period for each adsorption. As for EGA, it is known that this reagent destroys antigens of the Kell system. There is no reason why such RBCs cannot be used for adsorption, but the rate of autoantibody removal may not be as good as it is with ZZAP-treated RBCs, given the absence any proteolytic enzyme treatment to enhance the adsorption process.
ADDENDA Nov. 22, 2004
- A transfusion medicine physician in Barcelona reports that in his experience the performance of PEG autologous adsorption is controversial. He is aware that several authors have had success with this method, while others cannot validate the method in their laboratory. His institution started using a PEG autologous adsorption procedure two years ago, and he reports that it has worked very well. They have made comparisons with their reference test (ZZAP adsorption) and they have had the same results with the two techniques. They perform PEG autologous adsorption whenever they have a positive direct antiglobulin test in a pretransfusion sample. He reports that their data for this method will be published in Transfusion in 2005. He states they have no experience with EDTA-glycine method for performing adsorption procedures. They use that method for phenotyping red cells with bound antibodies.
ADDENDA Jan. 9, 2005
- Editor's comment: Colleagues may find the prior e-Network Forum discussion Warm autoantibody-adsorption procedures with PEG to be of interest to the current dialogue.
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Ira A. Shulman, MD
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