A Tale of Two Types

“It was the best of times, it was the worst of times.” In the blood bank, some of the best days can come from some of the worst days. When we come together as a team to work on a puzzling antibody problem, or to respond to a trauma, we can take pride in our work and know we have done our best to help the patient. In the blood bank we are constantly being called upon to learn and to be “disease detectives.” These are the best times. I tell my students that antibody panels are like puzzles and ABO discrepancies are mysteries to solve. Of course, when the Emergency Room is calling for blood for a trauma, or the Operating Room has an emergency surgery on a patient not previously type and crossed, any “problem” to solve can be a bit stressful.

ABO discrepancies are one challenge we face in blood banking. These are generally not clinical problems, but are serologic problems encountered by the blood bank technologists. Some discrepancies are easier to resolve than others, but still usually require a bit of investigation, and time. We don’t see these every day, so they can set us back a step when we do come across them.

One such situation that I recall was a young man in the ER who arrived by ambulance after a motor cycle accident. My trauma beeper went off and I called the ER to see if they wanted blood right away. Typically in these cases we bring them O blood in a cooler, and continue to use type O until we have a blood sample and current type, (performed twice if no prior history) and an antibody screen. In this case we were fortunate in that we got a sample almost immediately, before they started any transfusions. The type and screen was put on our Provue, but the instrument flagged an error on the type. When looking at the gel card, I could see mixed field reactions. Serology results are shown.

Anti-A Anti-B Anti-D Rh cont A cells B cells ABO/Rh
2+mf 0 2+ 0 0 4+ ?

ABO discrepancies occur when unexpected reactions occur in the forward or reverse grouping or the forward typing does not match the reverse typing. In general, RBC and serum grouping reactions are very strong; therefore reactions less than 3+ usually represent the discrepancy. In this case, testing patient cells with anti-A gave a 2+ mixed field reaction and patient cells and anti-D was only a 2+ reaction. The first step was repeating the test with the same sample. The repeat tube typing gave the same results. Additional steps included testing a new sample, completing the antibody screen, which was negative, and reviewing the patient history. At this time, we did have a positive identification on the patient and a medical record number. The patient had no previous Blood Bank history. However, reviewing the ER admission notes, it was noted that the patient had received 2 units of O negative packed cells in the ambulance en route to the hospital. Viewing the anti-A and the anti-D tubes under the microscope confirmed presence of mixed field agglutination.

Mixed field agglutination describes the presence of two populations of red cells. Mixed field agglutination is seen as small or large agglutinates in a field of many unagglutinated cells. In this case, we observed mixed field agglutination with the patient’s own circulating type A positive red blood cells agglutinating with the anti-A antisera, and the type O donor cells he received remaining unagglutinated. Patients can show mixed field reactions after recent out of group transfusions of as few as 1 or 2 units of packed cells. As well, when group O packed RBCs are transfused to a group A, B or AB recipient, there is always a small amount of plasma transfused. Thus, anti-A, anti-B and anti-A,B are almost always passively transferred. Even though it is unlikely that the passively acquired ABO antibodies will cause in vivo hemolysis, it would be recommended to continue transfusing O blood instead of type specific blood for the duration of the immediate episode and until anti-A antibodies are no longer detectable in the patient’s serum.

This case is an example of an artificial chimerism. Chimerism is the presence of 2 cell populations in a single individual and, in this case, was easily explained by the recent out of group transfusions.  This patient was sent to surgery and continued receiving several more units of group O RBCs during and after surgery. The patient’s blood type continued to appear as a mixed cell population during his hospital admission.

There are a number of other scenarios in which mixed field reactions could cause a discrepancy in a patient’s ABO/Rh typing. Some weak subgroups of A (A3) are known for giving mixed field reactions. Mixed field reactions can also be seen in other artificial chimera cases, such as are seen with transplanted bone marrow or peripheral blood stem cells of a different blood type.  If mixed field reactions are present, review the patient’s transfusion history to determine if the patient has been transfused with non-group specific RBC components in the past 3 months or received an ABO-mismatched stem cell or bone marrow transplant. More uncommon and unusual are cases of true chimerism, which can occur with fraternal twins.  Stay tuned for my next transfusion medicine blog for a discussion of chimerism!

A few key tips to remember when encountering an ABO discrepancy:

  • Retest the sample first, using a different method, if available
  • Check for technical or clerical errors
  • Remember that the weakest reactions are usually the ones that are in doubt
  • Complete the antibody screen and note positive reactions
  • Check the patient diagnosis
  • Check Blood Bank history
  • Most of all, take a deep breath and relax. You can solve this!

References

  1. Charles Dickens. A Tale of Two Cities. 1859
  2. George Garratty. Problems Associated With Passively Transfused Blood Group Alloantibodies. AJCP, June 1998
  3. Denise M. Harmening, Modern Blood banking and Transfusion Practices, Sixth edition, 2012.
  4. Christopher Sharpe, et al. Mixed field reactions in ABO and Rh typing chimerism likely resulting from twin haematopoiesis. Blood Transfus. 2014 Oct; 12(4): 608–610.

Socha-small

-Becky Socha, MS, MLS(ASCP)CM BB CM graduated from Merrimack College in N. Andover, Massachusetts with a BS in Medical Technology and completed her MS in Clinical Laboratory Sciences at the University of Massachusetts, Lowell. She has worked as a Medical Technologist for over 30 years. She’s worked in all areas of the clinical laboratory, but has a special interest in Hematology and Blood Banking. When she’s not busy being a mad scientist, she can be found outside riding her bicycle.

Blood Bank Case Study: Once Upon a Discrepancy

I have taught Transfusion Medicine to MLS students for a number of years, and one of the more challenging concepts for my students is that of ABO discrepancies. We use ‘dry’ labs for ABO discrepancy examples because it would be difficult to create actual samples that illustrate the various scenarios. Without seeing this in the lab, and actually performing the steps to resolve, visual learners in particular can be at a disadvantage. In reality, some of the more unusual ABO discrepancy problems are found more often on exams than in real life. Consequently, in the Blood Bank lab, when a technologist comes upon an ABO discrepancy, it can be something they are not very experienced with and it can be more scary than exciting. I have always felt that one of the best things about being a medical technologist is that we get to solve puzzles and find answers. So, let’s put on our detective hats and follow along with our case history story of an ABO typing discrepancy.

Once upon a discrepancy… a forward typing did not match a back typing. The first thing the tech did was to repeat the typing. Many labs recommend using a different method in the repeat, so if typings are routinely done by an automated method, a repeat testing might be done by tube typing. In this case, we can see the results of the initial testing and the results of the tube typing below:

Automated typing

Reagent Anti-A Anti-B Anti-D A1 Cells B Cells Interpretation
Results  4+  0  4+  1+  3+  ??

Repeat tube typing

Reagent Anti-A Anti-B Anti-D D Control A1 Cells B Cells Interpretation
Results  4+  0  4+  NT  1+  4+  ??

 

As you can see, the repeat typing simply rules out technical or clerical errors and confirmed that the testing was performed correctly. So far so good. However, since we got the same results on repeat testing, what is the next step in resolving this discrepancy?

I teach my students to think of a few ground rules when working on ABO discrepancy problems. The first is that, typically in these situations, it is the weak reaction that is the discrepant one. We have a patient who front types as an A, but the back type looks like an O. With ABO typing we usually get fairly strong reactions, so the 1+ reaction with A1 Cells is the suspect one. The second rule of thumb is that antibody problems are much more common than antigen problems. Having and extra antibody reaction or missing an antibody reaction is more common than extra or missing antigens. In this case we have an extra antibody reaction. This patient looks like a group A who is making anti-A1 which has reacted with our A1 cells.

Our next step is to discover why we have an extra antibody. I would like to emphasize the importance of looking up the patient’s history to help you resolve a discrepancy. This is the third thing that should always be done when investigating an ABO discrepancy. Accurate patient history including any previous Blood Bank results, age, pregnancy history, medications and diagnosis can all be used to help resolve these problems.

At this point techs are probably thinking ‘This is easy!’ and thinking about A subgroups. Remember that about 80% of group A people are group A1 and about 20% are group A2. There are also other less common subgroups of A, but A2 is the one that we encounter most often. Some group A2 people can make anti-A1, either naturally or as an immune response. This patient is a 30 year old woman who is in the Emergency Room and has just been scheduled for surgery. The physician has ordered a type and crossmatch for 2 units of blood. A look at her medical history shows she has never been pregnant nor has ever received blood products. We have no previous Blood bank history on the patient. While an anti-A1 can be from previous transfusions or pregnancy, it can also be naturally occurring. This seems to support our speculation that she is an A2 subgroup with a naturally occurring anti A1, so while we are waiting for our screen results, we perform A1 lectin testing. The results are shown below:

abo-disc

If our patient was group A2 as we thought, her A2 cells would not react with anti-A1 and her plasma would not have anti-A2 and would not react with A2 cells. Our results do not match our original hypothesis that the patient is group A2 and we can rule out a subgroup of A. What is her type, and what is causing the discrepancy?

To help solve this discrepancy, the tech looked at the solid phase screening results only to find that the screen was negative, thus making this puzzle even more perplexing. He repeated the screen in tube at IS, 37C and AHG and found positive reactions. Working up the panel, Anti-M was identified!

So, what type is this patient? She is group A1 pos with an cold reacting anti-M antibody. The policies of the medical center would determine if this patient should be given cross match compatible units that are not antigen typed or crossmatch compatible M negative units.

Anti-M is a naturally occurring cold antibody. Most examples of Anti-M are IgM, do not react at 37C and are not considered to be clinically significant. However, anti M can also present with an IgG component and react at 37C and AHG. In this case, it would be considered clinically significant and any units transfused must be negative for the M antigen.

This patient’s anti-M was only reacting at IS and determined to be not clinically significant. Despite this, we have seen that non-ABO alloantibodies can and do interfere with ABO typing and are a common cause of unexpected reactivity in ABO reverse typing. Performing the ABO testing at warm temperatures or repeating the reverse grouping with reagent A1 and B cells that are negative for M antigen can eliminate the cold reactivity and help resolve the discrepancy. It is important to remember that we must not only recognize discrepant results, but also resolve them adequately. Correct blood typing of patients is essential to prevent ABO incompatible transfusions and to help prevent alloimmunization.

References

  1. http://www.haabb.org/images/14_Hamilton-Neg_Ab_screen_For_website.pdf
  2. Harmening, Denise M. Modern Blood banking and Transfusion Practices, 6th Ed. 2012
  3. Safoorah Khalid, Roelyn Dates, et al. Naturally occurring anti M complicating ABO grouping. Indian Journal of Pathology and Microbiology. Vol 54, Issue 1, 2011. P 170-172

 

Socha-small

-Becky Socha, MS, MLS(ASCP)CM BB CM graduated from Merrimack College in N. Andover, Massachusetts with a BS in Medical Technology and completed her MS in Clinical Laboratory Sciences at the University of Massachusetts, Lowell. She has worked as a Medical Technologist for over 30 years. She’s worked in all areas of the clinical laboratory, but has a special interest in Hematology and Blood Banking. When she’s not busy being a mad scientist, she can be found outside riding her bicycle.

Poll: Ficin-Treated Red Blood Cells

Read the paper in Lab Medicine or listen to the podcast to learn more about one institution’s ficin protocol.

 

 

Acquired B Phenotype

Students learning about the ABO blood group system commonly get confused about two unique situations: The Acquired B phenotype and the Bombay phenotype.

These two entities are VERY different, but they are similar in this way: people are asked about both on exams all the time, but hardly anyone every actually SEES either one in real life! It is essential for students of blood banking to understand Acquired B clearly, as it remains a real possibility in everyday practice. I’ll cover Acquired B in this month’s blog, and next month I will discuss Bombay.

Routine ABO testing is performed in two distinct (but usually simultaneous) stages, known as “red cell grouping” (forward grouping or “front type”) and “serum grouping” (reverse grouping or “back type”). Here’s an example of how it works: If a person’s red blood cells (RBCs) react strongly with reagent anti-A but not anti-B, we would interpret their red cell grouping as blood group A. If there is no ABO discrepancy, that same person’s serum should have no reaction with reagent group A1 RBCs and strong reaction with reagent group B RBCs (demonstrating the expected presence of anti-B in the serum). Thus, the serum grouping interpretation would also be blood group A, and no ABO discrepancy would exist (see this illustrated in the figure below).

aboexample

ABO discrepancies occur any time the interpretations of a person’s red cell and serum grouping do not agree. ABO discrepancy takes on many forms, and acquired B is a great, if not terribly common, example.

Students learning about the ABO blood group system commonly get confused about two unique situations: The Acquired B phenotype and the Bombay phenotype.

Usually, Acquired B occurs when the RBCs from a blood group A patient come in contact with bacterial enzymes known as “deacetylases.” These enzymes, commonly carried by bacteria that live in the colon, catalyze the removal of the acetyl group from the residue that gives the A antigen its specificity, N-acetylgalactosamine (GalNAc). This modification leaves the A-specific sugar as galactosamine (N-acetylgalactosamine with the acetyl group removed = galactosamine). Recall that normally, the group B-specific sugar is galactose.

acqb

As a result of this modification, anti-B in both human group A serum and especially certain monoclonal reagents will weakly agglutinate the group A RBCs carrying the acquired B antigen. This means that the patient’s RBCs may have a weakly positive reaction with anti-B in serum grouping tests instead of the expected negative (see image below). The serum grouping for these patients is no different from that expected for a group A individual (negative with group A reagent RBCs, strong positive with group B RBCs).

acqbabotesting

So, what does this actually mean? How do these patients actually get transfused? This is where the recognition of the entity in a transfusion service or reference laboratory is essential. Several simple strategies can be employed to prove that this patient is really NOT group AB. First, I always advise people to check the patient history! The rare cases of acquired B that are still seen will often be associated with colorectal malignancy, gastrointestinal obstruction, or gram-negative sepsis (where those bacteria can contact the RBCs). Second, adding the patient’s own serum to his RBCs (autoincubation) reveals no incompatibility. In other words, this patient’s own very strong anti-B does not recognize the acquired B antigen (which is really just a partially modified group A antigen) as being an actual group B antigen. We already know that this patient has anti-B in his serum from his serum grouping results (see above), but the patient’s own anti-B completely ignores the acquired B antigen on his RBCs (even though human anti-B from other people will react). Third, the technologist can use a different form of monoclonal anti-B in the patient’s red cell grouping test. Certain clones are known to react with acquired B, while others are not (normally specified in the package insert), and choosing a different clone (often easier in reference lab settings) will render the forward grouping consistent with that of a group A person. Also, incubating the Acquired B RBCs with acetic anhydride will lead to “re-acetylation” of the modified A antigen and loss of the B-like activity. Finally, acidifying the reaction mixture of the patient’s RBCs with human anti-B (non-self) can eliminate the incompatibility with that source of anti-B.

In the end, Acquired B is a serologic problem that is fairly easy to recognize, especially on examinations (I always tell my students that when they see a problem that starts with words like, “A 73 year old male with colon cancer…”, check the answer for Acquired B!). In real life, experienced blood bankers can diagnose and confirm Acquired B fairly easily in the rare times that it is seen. These patients can receive group A blood without a problem, and the ABO discrepancy will disappear as the infection or other situation causing causing contact with bacterial enzymes clears. Thanks for your time and attention. See you next month when I will discuss the Bombay Phenotype!

 

Chaffin

-Joe Chaffin, MD, is the new Vice President and Chief Medical Officer for LifeStream, a Southern California blood center headquartered in San Bernardino, CA. He has a long history of innovative educational efforts and is most widely known as the founder and chief author of “The Blood Bank Guy” website (www.bbguy.org).

Stocking Shelves

My struggle in the community hospital setting is having the appropriate inventory for the patient population I need to serve. When I stocked the refrigerator during my golf club days the oldest inventory went up front and the new product went to the back. Later in graduate school I learned that was the FIFO method of inventory management. Blood Bankers have a unique twist thrown our way in that as blood sits on our shelves certain things happen that make an older unit less desirable than one collected a few days prior. The life span of a red cell is around 100-120 days depending on which literature you cite. Our job as blood bankers is to get the freshest blood to each patient we serve, so inventory management becomes more of an art than science.

Let’s take first the type specific debate. Some will say always transfuse type specific blood; if the patient is type A then the patient receives type A blood. Some will say to give whatever is most fresh; if we have fresh O cells an A person will get O. What I found when I first came to be the supervisor in my blood bank is that we were outdating a lot of type A blood. So instead of just decreasing the amount of type A, I also increased the number of type O I had on my shelf. This allowed me to be more flexible; I would give out more O when my inventory of A was low. Also, the blood I was giving out was always fresher than before I changed the inventory.

Let’s take this another direction. My policy states that any patient with an antibody has to have two red cell units set up so there is no delay if a transfusion is necessary. I would rather have two type O units typed for some antigens, because if the patient with the antibody doesn’t need it, the units are readily available to anyone else.  I use the flexibility of type O blood to be more versatile and to make sure that my patients are getting the freshest possible unit. I have searched for literature that says giving type specific blood is better for patient outcomes but I haven’t found it. If anyone has literature on the topic please send it my way.

This really comes down to what type of setting your blood bank serves. If you are in a medium size community hospital you will need to make these type of decisions to be flexible with your inventory. If you are a large medical center and are going through blood as soon as it gets delivered then you may not have to worry as much. The majority of us do not work for large centers, however, so we must look and analyze how we can best use this precious resource.

 

TommyTransfusion

Tommy Transfusion is the pseudonym of a blood bank supervisor in the midwest.