Order of the Steps in Serologic Testing

This is a teaching document I give to new staff, medical technology students, pathology and residents.  Very often I get the question, “Why can’t I just do the antiglobulin phase crossmatch first and then phenotype the RBC unit?” Or:  “Why do I have to add reagents in a particular order?”

My practice has always been to select an antigen-negative RBC unit first, then do the antiglobulin-phase AHG crossmatch.  This way I know that the unit was definitely phenotyped before release.  Likewise, the blood bank computer now only offers antigen-negative units for allocation and then crossmatching if there is a clinically significant antibody.

In a manual setting without a blood bank computer system, performing the AHG crossmatch may yield a negative result, even if the unit is antigen-positive.  With storage, some antigenic expression is weakened so it may not be detected at the time of crossmatch.  Yet, there may still be enough antigen present to cause hemolysis.  Not detected does not necessarily mean not present!!

I expect that many inexperienced staff may be tempted to forego the antigen typing if the AHG crossmatch is negative.

This is an analogous logic to the question, “Do I add the cells or the antiserum/plasma/serum first for the reaction?”  If you add the cells first, you may forget to add the patient’s plasma/serum or a typing antiserum and you might not be able to detect the omission by looking at the tube or gel.  Actually, I once recommended to one vendor that it color the typing antiserum so it was conspicuously showing on the gel.

I was taught that this is a matter of discipline to ensure that all steps are performed.

However, for every practice, there has to be flexibility.  If there is no typing reagent or if it is very expensive or in short supply, one may have no choice but to do the AHG crossmatching first.  Often there is still another option:  one can often preliminarily screen units first before using a rare reagent—examples:

  • Check if patient is group O first and antibody screen panreactive in suspected anti-H.
  • Check P1 typing first if there is a suspected anti-PP1Pk.
  • Check the antibody screen for panreactivity first for antibodies of high-incidence or prevalence antigens.

ABO Incompatible Stem Cell Transplant: What ABO type of RBCs Should I Use?

In an ABO-incompatible stem cell transplant, both donor and recipient RBC types may be present.  Likewise, immune effector cells from both the donor and recipient may be present.

Using group O RBCs and AB plasma is an option but there are limited supplies of both.  Since we use RBCs in additive solution (SAGM), only minimal residual donor plasma is available and unlikely to be clinically significant.

Here is my approach:

  1. Use fresh specimen to perform forward and reverse type
  2. Check the reverse type:  does it show either anti-A and/or anti-?
    1. If anti-A detected, do not transfuse group A RBCs.
    2. If anti-B detected, do not transfuse group B RBCs.
    3. If both anti-A and anti-B detected, continue using only group O RBCs.

Massive Transfusion: When to Revert to the Original ABO Type

In a massive transfusion setting, the patient’s forward and reverse type will reflect the use of group O RBCs and the patient may even fully type as group O, depending on the number of units transfused.  Group O RBCs are very precious and in short supply so we need to switch the patient back to his/her own type as soon as feasible.  RBCs in additive solution (e.g. SAGM) have only minimal residual plasma so the load of anti-A,B from the O cells is minimal.  In my organization, we did not titer ABO hemolysins in blood donors.

Here is my approach:

  1. Use fresh specimen to perform forward and reverse type
  2. Check the reverse type:  does it show either anti-A and/or anti-B from the group O massive transfusion?
    1. If anti-A detected, do not give group A RBCs.
    2. If anti-B detected, do not give group B RBCs.
    3. If both anti-A and anti-B detected, continue using only group O RBCs.

Some people would recommend performing a full AHG crossmatch using the patient’s current plasma and RBCs of the original ABO type:

  1. If compatible, return to the original ABO type.
  2. Otherwise, continue using group O RBCs.

I did NOT do the full AHG crossmatch and had no hemolytic transfusion reactions if the RBCs were compatible with the current reverse type plasma.

Antibodies to Public Antigens: NGHA Experience

This is an update of a presentation I gave while working as Head of Transfusion Medicine for Saudi Arabian National Guard Affairs, King Abdulaziz Medical City at Riyadh. I used this strategy there and later at HMC Doha.

Fortunately we had access to rare antisera such as anti-Tja (anti-PP1Pk) which we could purchase from Diamed AG and its successor Biorad. I emphasize that we only sparingly used these rare antisera AFTER ruling out more common high-incidence antibodies.

Teaching Document: Irradiating RBCs from Outside Transfusion Medicine

This is a teaching document of a process to irradiate RBC components when our irradiators are not functioning.  In our system, we used Mirasol pathogen-inactivation so our RBC units were affected.  The number of units for advance irradiation was based on our historical usage of irradiated units across our system.  The workweek is Sunday-Thursday so on Thursday special effort was made to have the minimum number of irradiated units available.

We used irradiated units for compatibility testing to avoid the possibility that the unit would be released without irradiation.

Emergency Interim Procedure:

  1. Verify the patient’s diagnosis/location:  All Hematology-Oncology patients should receive irradiated blood.  Refer to the list of diagnoses for which irradiation is indicated (attached).
  2. Only PRBCs need to be irradiated.  Mirasol-treated (pathogen-inactivated) platelets can be used directly without irradiation in accordance to Council of Europe CE regulations.
  3. Attach radiation indicator labels to the selected units as per the irradiation procedure.
  4. Send the units for irradiation to the Radiation Oncology department.
  5.  Verify that the proper dose of irradiation was received while in the Radiation-Oncology department.
  6. Keep a minimum stock of 20 group O-positive units irradiated at the start of each day AND before the start of the weekend on Thursday afternoon.  Irradiate that number of group O-positive units plus any other specific requests for blood.
  7. You should use the irradiated units for crossmatching.  DO NOT CROSSMATCH FIRST BEFORE IRRADIATING!!  If the antibody screen is positive, refer the specimen to the Transfusion Service for further processing and selection of units.
    1. Note:  If a special antigen typing is needed, we may have to irradiate after selecting the antigen-matched unit if it is not found in the group of irradiated units.
  8. If a unit is required for a child < 20 kg, it must be washed before release if the unit was irradiated more than 24 hours previously according to our irradiation policy.