Resolution of discrepancies caused by anti-A₁

First step: We must determine if the person is group A₁ or group A₂. (If group A₁, the discrepancy with the A₁ cells is NOT due to anti-A₁). To do this, we antigen type the person's red cells with the anti-A₁ lectin which is Dolichos biflorus . Another source of anti-A₁ is "absorbed anti-A" which is made by absorbing anti-A with A₂ red cells.] If the red cells agglutinate, the person is group A₁. If the red cells do not agglutinate, the person is not group A₁, and probably is group A₂ assuming the red cells reacted strongly (3+ or 4+ with anti-A).

NOTE: Positive and negative antisera controls are set up with the antigen typing to control the reactivity and specificity, respectively.

Second step: If the person appears to be group A₂, we must prove that the extra antibody is anti-A, and not some other IgM irregular antibody. To do this we test the person's serum against a panel of 3 A₁ cells and 3 A₂ cells. If anti-A₁ is present, only the A₁ cells should agglutinate.

NOTE: 3 A₁ and 3 A₂ cells are required in order to ensure that the antibody reacting is anti-A₁ and not some other antibody. With 3 cells of each group we can achieve a statistical probability of 95% that the right antibody has been identified. For example, if only one A₁ cell and one A₂ cell were tested, by chance, another antibody like anti-M or anti-P₁ could react with the A₁ cells (if they were M+ or P₁+), but not the A₂ cells (if they were M- or P₁-).

Quality Control of second step

1. Do the tests in saline at RT or lower (e.g., 4°C) because anti-A₁ is an IgM antibody that reacts best in the cold.
2. Include an autocontrol: an autoanti-I may be reacting with the A₁ and A₂ cells as they are adult cells and I-positive. An autocontrol will reveal if this is happening.

Optional QC

Some labs believe it is more time-efficient to include more cells in the mini panel used to identify anti-A₁. Other cells that may be helpful are the following:

1. Cord red cells: an autoanti-I will react weakly or negative with cord cells because the I antigen is very poorly developed at birth.
2. Group O screen cells: these will detect whether or not an irregular IgM antibody like anti-M or anti-Le^a is agglutinating the A₁ or B cells.