Transfusion Medicine Case Study: Positive Pretransfusion Test

A 72 year old man was admitted to the hospital for an aneurysm repair. The physician ordered a type and crossmatch for 6 units of blood in preparation for surgery. The patient history included surgery in 2016 during which he was transfused with 4 units of RBCs.

patient’s blood type: A positive

antibody screen: negative

history: anti Jkb  (2016)

6 Jkb negative units were found and full crossmatches were performed. One of the 6 donor units was incompatible. What is the most probable explanation for these findings?

If the patient has a negative antibody screen, and no history of an antibody, most facilities would do an electronic crossmatch or an immediate spin crossmatch. The immediate spin (abbreviated) crossmatch will simply verify ABO compatibility. However, if the patient has a positive antibody screen, we must identify the antibody, phenotype the patient, and do a full AHG crossmatch with donor units that are antigen negative for the corresponding antibody. In this case, the patient had a history of an antibody, so the antibody must be honored, and antigen negative units must be chosen for transfusion.

Kidd antibodies demonstrate dosage, are often weak, and can be found in combination with other antibodies. Because if this, they can be notoriously difficult to detect. They are usually IgG and are made in response to transfusion or pregnancy. Jkb has an antigen frequency of about 73% in the white population and about 43% in the black population. To find antigen negative blood, we consider that about 27% of units would be antigen negative. The tech working on the sample screened 21 units and found 6 that were Jkb negative.

AHG crossmatch results:

unit 1: compatible

unit 2: compatible

unit 3: compatible

unit 4: 3+ at AHG

unit 5: compatible

unit 6: compatible

There are 2 possible scenarios for the above results. A crossmatch is a test between donor’s red blood cells and patient’s plasma. Antigens, we know, are on red blood cells and antibodies are detected in the plasma. So, with a negative antibody screen, crossmatch incompatibility is due either to a patient antibody to a low incidence antigen on the donor red blood cells, or a donor cells with a positive direct antiglobulin test. We can easily rule in or out a positive donor DAT by performing a DAT on the segment. If the donor unit has a positive DAT, the unit should be quarantined and the positive DAT reported to the collecting facility. If the donor unit has a negative DAT, the patient likely has an antibody to a low incidence antigen.

Low frequency antigens are uncommon, but antibodies that recognize them are less rare. Fortunately, for patients with these antibodies to low frequency antigens, finding antigen negative compatible blood is easy. As we can see, 5 of the 6 chosen units were negative for the unknown low frequency antigen and were antiglobulin crossmatch compatible. The low prevalence of the antigen makes compatible blood readily available. If transfusion is necessary, it should not be delayed while waiting for identification of the antibody.

In this case, the antibody screen was repeated and the negative result was verified. In many cases, it may not be possible for a lab to identify the antibody because the lab may not have the necessary panel cells or typing reagents. Yet, these antibodies to low incidence antigens that react at AHG can be clinically significant and cause severe hemolytic transfusion reactions. To identify the antibody, you may need to send the sample to a reference lab for testing against a panel of reagent red cells that express low incidence antigens. Alternately, the donor red cells that were incompatible can be tested against known antibodies to low prevalence antigens  to help identify the antibody.

In this patient, anti-Wra was identified. The incompatible donor unit was verified to be Wra positive. Wra is part of the Diego system, usually IgG, and has ben implicated in hemolytic transfusion reactions.

One of the reasons I have written up this case is questions my Transfusion Medicine students often ask about exam and exam prep questions concerning incompatibility. Below are 2 questions to give examples of the confusion.

“At the indirect antiglobulin phase of testing, there is no agglutination between patient serum and screening cells. One of 3 donor units was incompatible.. The most probable explanation for these findings is that the:

  1. patient has an antibody directed against a high incidence antigen
  2. patient has an antibody directed against a low incidence antigen
  3. donor has an antibody directed against donor cells
  4. donor has a positive antibody screen”5

answer: b

“Which of the following would most likely be responsible for an incompatible antiglobulin crossmatch?

  1. recipient’s red cells possess a low incidence antigen
  2. anti-K antibody in donor serum
  3. recipient’s red cells are polyagglutinable
  4. donor red cells have a positive direct antiglobulin test”4

answer: d

I am asked why is one answer “low prevalence antigen” and one answer “positive DAT”? I typically ask questions of my students to let them reason out the answer. Take a careful look at the words antigen and antibody. Remember that a DAT is a test of red cells, the IAT tests for antibodies in plasma. A crossmatch uses donor red cells against patient plasma. Therefore, even though these are both reasons for the incompatibility of one out of multiple units, each question only has one answer of a common reason for such incompatibility. Be sure to read questions and use your theory and knowledge of testing when encountering discrepancies and problems in Blood Bank. To all of my students: Happy Studying for your ASCP exam!

References

  1. Fung, Mark K., Technical Manual 18th ed, Bethesda: AABB, 2014.
  2. Harmening, Denise M. Modern Blood Banking and Transfusion Practices, 7th edition, 2019.
  3. Schonewille, Henk, et al. “The importance of antibodies against low‐incidence RBC antigens in complete and abbreviated cross‐matching”. The Journal of AABB. 20 June 2003.
  4. BOC Study Guide, 5th edition. Clinical Laboratory Certification Examinations.  ASCP, 2016
  5. BOC Study Guide, 6th edition. Clinical Laboratory Certification Examinations.  ASCP, 2018

-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.

The Best Gift of the Season: A Gift of Self

We few, we happy few, we band of brothers;

For he today that sheds his blood with me

Shall be my brother.

Henry V (Act 4, scene 3), Shakespeare.

Good morning! We’re entering the holiday season, and it’s an exciting time for all. I love seeing the ethnic and cultural diversity as we all celebrate our favorite holidays with family and friends. I myself look forward to the holiday season. It’s a festive time and a season of giving and sharing. It’s a favorite time of year to share traditions and create new ones. However, at a time when stores have Christmas candy on the shelves, holiday lights up and holiday music playing on the day after Halloween, I feel a bit rushed and want to slow down and find better ways to celebrate and enjoy the season. Over the past few years I have been making a special effort to become more environmentally conscious; remembering my reusable bags at stores, purchasing more reusable products, and reusing, recycling, and upcycling whenever I can. I belong to a community ‘buy nothing’ group and am warmed by the generosity of strangers to others in the community. It’s wonderful to give from our abundance and to receive wish list items from neighbors without having to exchange money. And it’s great for the environment, too. Used items are being put to use by others, and not into landfills. People in the community have asked for or gifted furniture, clothing, tools, toys and many other goods and services. I have gifted no longer needed clothing, household items, excess fabric from my fabric stash, and donated my time to participate in a career fair at a local high school. I have been given a car set for my grandchildren when they visit, toys, and someone even loaned me a bike trailer so we could take my granddaughter out for a bike ride. The generosity makes it feel like the holiday season all year round.

So, you may be asking, “where is this blog going?” I saw a memo from Red Cross this week that there is a critical need for blood and platelets and thought that giving to our community with the gift of blood would be a wonderful way to make this holiday season even better! It’s one of the most generous gifts we can give, and costs nothing. Every 2 seconds in the US, someone needs a blood product. That’s about 36,000 units of red blood cells, 7,000 units of platelets and 10,000 units of plasma needed every day. 21 million blood products are transfused every year.1 That’s a lot of blood. And, these blood products cannot be manufactured, so must come from volunteer donors.

In the US, we need to collect about 13,000 units a day to meet demand. Approximately 14 million units of whole blood are collected each year from roughly 7 million donors.1 The blood is processed into components and used in the treatment of surgical, obstetric, oncology, and other patients. One unit of whole blood can be made into up to 3 components and used to help up to 3 patients. Yet, even with all these donations we still cannot keep up with demand. Weather, holidays, illness and travel can all affect blood donations. Shortages are not just apparent during the winter holiday season. This past summer, the Red Cross announced a critical blood shortage around the July 4th holiday. Compared to other weeks, there were 17,000 fewer blood donations during the week of July 4th. As of July 9, the Red Cross had less than a three-day supply of most blood types and less than a two-day supply of Type O blood. 2 During the summer, and particularly during the holiday week, people are busy with other activities or traveling. In the winter, busy schedules, holiday travel, winter weather and seasonal illnesses contribute to fewer blood and platelet donations. Severe weather can also cause the cancellation of blood drives which greatly impact the blood supply.

Some people donate blood because they see this critical need and hear the calls for blood. Others donate because a classmate or friend asked them to. Some people feel it’s their civic duty. For some, it just makes them feel good to help another person. And, others donate for the cookies and tee shirt. Yet, for all donors, it is a form of volunteerism and giving to the community. But, did you know that, other than the benefits from helping others, there are benefits to the donor, as well? Helping others can improve our emotional and physical health. It can help reduce stress, improve emotional well-being and help people feel a sense of belonging. A study conducted in Sweden concluded that regular blood donors enjoy better than average health.Blood donors had an overall mortality 30% lower and a cancer incidence 4% lower than the control population.3 Donating blood may help reduce high iron stores, a risk factor for heart attack. In addition, there have been several studies over the past few years, exploring the hypothesis that regular blood donations may help in the management of hypertension and high cholesterol.

Another interesting benefit of blood donation is being able to contribute to science and research. For example, there is currently a study being conducted on donor blood to test an investigational nucleic acid test for Babesia microti. Babesia microti is responsible for most transfusion-transmitted babesiosis cases in the United States, but there is no licensed test for screening for B. microti in donated blood. Participation in this study can help obtain FDA approval for a screening test. By giving your consent to use your blood sample, there is no additional blood taken and no further time commitment, but you can help protect the public health by supporting the development of a new blood safety test.

How can we, as individuals, help? About 38% of the population is eligible to donate blood, but less than 10% of the population actually donates. To be eligible to donate, you should be in good general health and feeling well. You must be at least 17 years old in most states (16 years old with parental consent in some states) but there is no age limit to donation. Adult doors must weigh 110 lbs, but there are additional height and weight requirements for donors 18 years old and younger. There have also been some recent changes to blood donor requirements. I will not be able list all of them here, but some of them don’t change a deferral, only the reasoning behind the deferral. One of the most prominent changes is, as of 2016, the indefinite deferral for men who have had sex with men, has been changed to a 12 month deferral since the last sexual contact with another man . Also changed is the minimum hemoglobin for male donors. This has been raised from 12.5g/dl to 13.0 g/dl. Until this time, the cutoff was the same for both males and females. Males with a Hgb below 13.0 g/dl are considered anemic and are no longer eligible to donate blood. On the other hand, the criteria for females to be mildly anemic is a Hgb below 12.0 g/dl, so females between 12.0 g/dl and 12.5 g/dl, though not considered anemic, are still not eligible to donate. The minimum hemoglobin for females has not changed and remains 12.5 g/dl. To review other eligibility requirements, visit https://www.redcrossblood.org/donate-blood/how-to-donate/common-concerns/first-time-donors.html

So, in this busy season, we often find ourselves with little time to get our own “to do” lists done, yet alone volunteer our time for others. But most of us would welcome an hour to reduce stress and improve our emotional well-being. Please consider a gift of self this season. It takes about an hour of your time, you get to sit and relax with your feet up, to feel good about yourself, and you’ll even get a snack!

Happy Holidays!

References

  1. redcrossblood.org
  2. https://news.azpm.org/p/news-splash/2019/7/19/155196-fourth-of-july-donation-slowdown-leads-to-blood-shortage/
  3. Edgre, G et al. Improving health profile of blood donors as a consequence of transfusion safety efforts. Transfusion. 2007 Nov;47(11):2017-24.
  4. Kamhieh-Milz S, et al.Regular blood donation may help in the management of hypertension: an observational study on 292 blood donors. Transfusion. 2016 Mar;56(3):637-44. doi: 10.1111/trf.13428. Epub 2015 Dec 8.

-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: “Oh I’m so confused! What type am I?” Weak D Phenotypes in Pregnant Women

In my July post, “Blood Bank Case Study: What’s Your Type?” I discussed some of the dilemmas when dealing with a weak D phenotype and the fact that there is no standard or general consensus as to the testing performed or terminology to be used in resulting a weak D patient. Results obtained on patient testing also vary depending on the method used, and the anti-D reagent and enhancement used in testing. This can be confusing to medical technologists, physicians and to patients.

For anyone who has not been in the Blood bank for a while, the Du variant was first recognized in 1946 and renamed weak D in 1992. To review last month’s blog, serologic studies have distinguished three broad categories of D variants, weak D, partial D, and DEL, from conventional D. A serologic weak D phenotype is one that has no or weak reactivity (≤2+) of RBCs with an anti‐D reagent at immediate spin, but does agglutinate with antihuman globulin. Since there is no general consensus on how labs perform and report patient testing for weak D, it is left up to individual interpretation as to what type blood these patients should receive, and, if pregnant, if they should receive Rh D immune globulin (Rhogam). Last month I focused on testing, resulting, donors and blood administration. In this blog I will focus on issues concerning weak D in the obstetric population and how labs can move forward now and in the future towards the best patient care and blood management.

About 15% of Caucasians are RhD negative. About 3% are weak D phenotypes.In the genral population, this means that about 0.2% to 1.0% inherit RHD genes that code for serologic weak D phenotypes.2 In Europe and the US, weak D phenotypes are the most common D variants found, but we also know that the prevalence of weak D phenotypes varies by race and ethnicity. Today we have much more information about D antigen expression than we had in the past, because we have the availability to genotype these weak D RBCs. We know that more than 84 weak D types have been identified, but types 1, 2, and 3 account for more than 90% of these in people of European ethnicity.1  Currently, with the mixed ethnicity population in the US, about 80% of people who inherit RHD genes for serologic weak D phenotypes are found to be weak D type 1, 2, or 3.3 We also know that types 1, 2 and 3 are unlikely to become alloimmunized to anti-D, so they can safely be treated as RhD positive and receive RhD positive units.

The introduction of RhD immune globulin in 1968 is one of the great success stories in obstetrics. Rhogam has been used very successfully in developed countries in the prevention and treatment of hemolytic disease of the fetus and newborn due to RhD alloimmunization. The routine recommendation is that women who are candidates for Rhogam receive one dose at approximately 28 weeks’ gestation and a second dose after the delivery of an Rh pos baby. Additional recommendations are for administration of Rhogam after threatened miscarriage, abdominal trauma during pregnancy and before invasive diagnostic procedures.

But, who is a candidate? Any unsensitized woman who is RhD negative and who may be carrying or who delivers an RhD positive baby is a candidate for Rhogam. And, that brings us back to the problem that we have no standardization for the reporting of serological weak D phenotypes.

As an example, let’s look at a patient who has 3 children. Many labs do not do weak D testing on patients and report anyone who is RhD negative at immediate spin as RhD negative. This patient was typed at such a lab (Lab #1) as RhD negative, and received Rhogam for her first pregnancy. During her 2nd pregnancy, she had moved to a different state, and went to another lab (Lab #2) for prenatal testing. This lab performed serologic weak D testing and found this patient to be weak D positive and reported her type as RhD positive. Rhogam was not further discussed during this pregnancy and the patient did not receive Rhogam. The patient had blood drawn during her 3rd pregnancy at yet a third hospital (Lab #3). Some labs distinguish women who are pregnant or of childbearing age from the general population, and have different procedures on the reporting of RhD type on these women. This hospital’s procedure was to do weak D testing on all patients, but, in women of childbearing age, if weak D positive, they report these women as RhD negative. The patient was told she was RhD negative and would be a candidate for Rhogam. At this time the woman thought she remembered that she didn’t get Rhogam with her second pregnancy and was a little confused, but with 2 young children and pregnant with her 3rd, she simply followed the doctor’s recommendation and didn’t question further. When her 3rd child was 4 months old, she attended a Red Cross blood drive at work and donated a unit of blood. Soon she received a blood donor card in the mail that said she was RhD positive. At this point she was thoroughly confused and questioned all the lab results she had had done over the past 6 years. On her next visit to the doctor she questioned her obstetrician. The obstetrician recommended RhD genotyping. The woman was found to be weak D type 2. The doctor explained to her that all blood donors who are weak D are treated as RhD positive, but, that as a patient, policies and procedures vary. However, he also informed her that now that they had her genotype, she would be considered RhD positive. He explained that the genotype was DNA testing, would not need to be repeated, she would not need Rhogam for any future pregnancies and she could safely receive RhD positive blood products.

The American College of Obstetricians and Gynecologists (ACOG) guidance practice bulletin of 1981 recommended that recommended that RhD‐negative women “whether Du positive or Du negative” were candidates for Rhogam. Shortly afterwards, that recommendation was reversed and revised to read “[a] woman who is genetically Du‐positive is Rh‐positive and administration of Rh immune globulin is unnecessary.1 This remained the recommendation of the group until the latest version of this publication in 2017. The 2017 ACOG guidelines recommend giving Rhogam to weak D positive patients, “in appropriate clinical situations, until further studies are available.”3 Another comparative study published in 2018 reported inconsistency between national groups over how to treat weak D phenotypes and recommended the creation of international guidelines.4

Thus, the controversy over whether a pregnant woman who is weak D positive is RHD positive or RHD negative continues. The latest recommendations, and those of ACOG, are for a move to genotyping patients with a serological weak D phenotype. There are several benefits to this. As we can see from my case study example, genotyping put this woman at ease and gave her definitive answers about her blood type. It also can do the same thing for medical technologists and physicians. RHD genotyping only needs to be performed once on a patient. If performed at the first prenatal appointment, this would alleviate much confusion as to procedures and how to report the results. I have in blood bank, that whenever we have a weak D on a prenatal patient, there are questions about how to result them, and we refer to the SOPs. We also occasionally get a patient who had previously been typed elsewhere where the reporting procedures were different and there is therefore an apparent discrepancy between the current and historical typing. This causes frequent phone calls from physicians and nurses asking for clarifications on weak D types, and questions about Rhogam. Lastly, RHD genotyping could avoid confusion which could lead to transcription and computer entry errors when entering types on these patients. RHD genotyping would solve all of these problems and eliminate confusion.

Additional benefits of RHD genotyping are, if RHD genotyping was performed on all weak D transfusion recipients, we could save as many as 47,700 units of RHD negative RBCs annually.3 With the availabilityof molecular testing, there is no reason to administer RhD negative units to patients who can use RhD positive units. This could help alleviate the constant shortage of RHD negative units. With RHD molecular testing, these critical units could be reserved for patients who are truly RhD negative.

It may not be feasible for all laboratories to perform molecular testing for RHD genotypes, but reference laboratories should offer affordable testing for the most prevalent and clinically relevant RHD genotypes. From a study done of over 3100 laboratories, it was found that, at this time in the US, most labs are managing weak D phenotypes as RhD negative. Laboratories not performing weak D testing are essentially avoiding their detection. Clinical laboratories should instead increase the detection of serological weak D and interpret these with the use of RHD genotyping. Rhogam shortages exists, and RHD genotyping could save thousands of injections of Rhogam annually in the US alone, and at the same time, avoid the unnecessary administration of products to patients. The work group study calculated that annually, approximately 24,700 doses of unnecessary Rhogam could be avoided.1 It is time to move forwards to molecular testing for the best patient care and blood management.

References

  1. Sandler SG, Flegel, WA, Westhoff CM, et al. It’s time to phase in RHD genotyping for patients with a serologic weak D phenotype. Transfusion 2015;55:680‐9
  2. Garratty G. Do we need to be more concerned about weak D antigens? Transfusion 2005;45:1547‐1551.
  3. Practice Bulletin No. 181: Prevention of Rh D AlloimmunizationObstetrics & Gynecology: August 2017 – Volume 130 – Issue 2 – p e57-e70 doi: 10.1097/AOG.0000000000002232
  4. Sperling, JD et al. Prevention of RhD Alloimmunization: A Comparison of Four National Guidelines. Am J Perinatol. 35(2):110-119. doi: 10.1055/s-0037-1606609. Epub 2017 Sep 14.

-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.