Complement-coated red cells for quality control of anti-C3d reactivity in an ID-MTS gel system for direct antiglobulin testing

A colleague at a transfusion service affiliated with the United States Armed Forces reports that their laboratory is in the process of implementing an ID-MTS gel system. A question has arisen as to what others consider a preferred approach for performing quality control on the MTS Anti-IgG,-C3d cards, which are licensed for direct antiglobulin testing. Although he understands that the QC of the anti-IgG reactivity of the MTS Anti-IgG,-C3d cards can be performed using IgG-coated reagent red cells, he wonders about how best to perform QC of the Anti-C3d reactivity of the MTS Anti-IgG,-C3d cards. According to the package insert, "It is recommended that each lot be tested each day of use with positive and negative samples ... the anticomplement reactivity of this reagent can be assessed by using complement-coated cells". Yet, when the inquiring colleague contacted the card manufacturer, he got the impression that they did not have a source of complement-coated reagent red cells for use in the QC of these cards.

The inquiring colleague is looking to find a supplier of complement-coated cells that would be appropriate for use in this QC. Can any colleague provide advice as to a source of such cells, and share their experience with how well these cells perform when used in this QC?

The following responses were received.

ADDENDA Jan. 20, 2004

1. The Editor is of the opinion that to be compliant with the Clinical Laboratory Improvement Act (CLIA) that the performance of the direct antiglobulin test (DAT) needs to be controlled by doing appropriate QC, each day that the DAT is done. Furthermore, the QC that is done needs to be performed in compliance with the manufacturer's package insert instructions. My interpretation of the available information is that the QC done for each lot of MTS Anti-IgG,-C3d cards include testing with known positive and negative red cell samples, to verify that both the anti-IgG and anti-C3d components of the anti-human globulin reagent are viable. Thus, it would seem reasonable to test each lot of MTS Anti-IgG,-C3d cards on each day of use with three different red cell samples: 1) an uncoated red blood cell, 2) an IgG-coated red blood cell, and 3) a complement-coated red blood cell (eg, a C3b- and/or C3d-coated red blood cell). In order to run these cells through the MTS cards, these red cells need to be made up in suspensions of approximately 0.8% in MTS Diluent 2. On the other hand, once the MTS Anti-IgG,-C3d card passes daily QC, use of IgG- or complement-coated red cells (so-called 'check cells') is not required (or even recommended) to confirm the validity of a negative DAT result when testing a patient sample. The reason that 'check cells' do not need to be used to confirm negative DATs when using the gel system is that the gel test utilizes a controlled centrifugation where cards spin at a 90 degree angle to the plane, forcing red cells into the microtube where they come in contact with the antiglobulin reagents and gel particles. Because of the centrifugation process and characteristics of the gel, less dense serum globulins that have not bound to the red cells are unable to travel thought the gel. This prevents neutralization of the antiglobulin reagent within the microtube, thereby eliminating both the need for washing and the use of IgG- or complement-coated red blood cells to verify each negative reaction.
   
   
2. A colleague at an academic medical center in Cleveland reports that complement-coated red cells can be purchased from Immucor (Phone: 800-829-2553). Ordering information is as follows: Item # 7928, Complement EC3b Control Cell. This reagent is manufactured by Gamma Biologicals.
   
   
3. W. John Judd, FIBMS, MIBiol, Professor of Immunohematology at the Department of Pathology, University of Michigan Medical Center (attribution used with permission) reports that at his institution they use C3d (rather than C3b) coated RBCs from Gamma for daily QC of anti-IgG, -C3d cards; the same RBCs that they use for daily QC of tube anti-C3 reagents. Review of their daily gel QC records reveals that these RBCs yield a 2+ to 3+ reaction. The C3b-coated RBCs that are available from Gamma are used primarily for validating tube tests that give negative DATs with reagents containing anti-C3.

ADDENDA Jan. 21, 2004

4. A colleague in Ohio wants to know if she can assume from this discussion that people are performing daily quality control on their IgG/C3d antiglobulin tube reagent using both an IgG-coated cell and a complement-coated cell? She has worked in several institutions and has never seen that practice in place before, nor is that currently their local practice. They QC their IgG/C3d antiglobulin reagent with an IgG-coated cell and do not perform any QC of that reagent for its complement reactivity.
   
   
5. Editor's NOTE: The Ohio colleague assumes correctly that many institutions do QC their polyspecific direct antiglobulin test with both IgG coated (check) cells and with complement-coated cells (see the 1999 discussion). It is true that if a laboratory is doing a 'polyspecific' direct antiglobulin test by a TUBE method, and if the package insert for that tube test allows for daily QC to be done with non-reactive cells (negative control) and with IgG-coated (check) cells (positive control), that would be acceptable practice. However, the package insert for the GEL method polyspecific direct antiglobulin test using the MTS IgG-C3d Gel cards suggests using a negative control, a positive control for the IgG reactivity, and a positive control for the complement reactivity. The exact wording is as follows: "Quality Control - To confirm the specificity and reactivity of the MTS Anti-IgG,C3d Card, it is recommended that each lot be tested each day of use with known positive and negative samples. Reactivity must be present with the positive sample only. The anticomplement reactivity of this reagent can be assessed by using complement-coated cells"

ADDENDA Jan. 23, 2004

6. A colleague in Spain reports that his blood bank has been using IgG-coated cells and C3d-coated cells (provided by Immucor) to perform quality control (QC) for ID-MTS gel cards since 1993. In their experience, the IgG-coated cells give a stable 3+ to 4+ reaction in the microtube, and this reactivity can be observed over the entire usable shelf life of the IgG-coated cell reagent.. However, they find that the C3d-coated cells give 2+ to 3+ reactions during QC at the beginning of the QC reagent's usable shelf life, but over time this reactivity slowly decays, and towards the end of the QC reagent's useful life, negative results can even be seen. This loss of QC reagent reactivity with C3d-coated cells can be a real problem, according to the Spanish blood banker, especially if the next shipment of QC reagent does not arrive in time.

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