The Occurrence of Lewis Antibodies in Pregnancy

A 36 year old woman presented to the delivery room at a local county hospital at 39 weeks’ gestation. The doctor ordered a type and screen on the patient, the blood was drawn and sent to the Blood Bank lab. The Blood Bank tech looked up the patient’s Blood Bank history and noted that an antibody screen done at 28 weeks was positive, with an anti-Lea identified. The Blood Bank’s policy is to have 2 units of blood available for any patient with an antibody. As the Blood Bank tech was working on the sample, the physician sent a STAT order for 2 units RBCs for intrapartum hemorrhage.

Are Lewis antibodies clinically significant? AABB defines a clinically significant antibody as one that causes decreased red blood cell survival of transfused cells, one that causes hemolytic transfusion reaction or one that causes Hemolytic Disease of the Fetus and Newborn (HDFN).3 In the Blood Bank, we would always be cognizant of all three criteria, but in this case, we are particularly concerned with HDFN.

The Lewis system is of great interest in immunohematology because of its unique characteristics. The Lewis blood group system is the only one where the antigens are not produced by the red blood cell itself. We learn in immunohematology that red cell antigens are structures that are usually formed on red blood cell membranes, but Lewis stands alone in that the antigens are glycolipids that are formed in the plasma and then passively absorbed onto the red blood cell membrane. This forms a loose attachment and these antibodies can shed or elute off the RBCs in certain circumstances.

Because Lewis antigens are not formed on RBCs, Lewis antigens are not present at birth and therefore not found on cord blood cells. Cord blood and RBCs from newborns will phenotype as Le(a-b-). The saliva of these newborns will have Lea and/or Leb antigens depending on the genes inherited, but the RBCs will test negative for these antigens at birth. By about 10 days of age, the Lewis antigens can be detected in plasma, and they will shortly thereafter begin to be absorbed onto the RBCs. Yet, children do not exhibit their true Lewis phenotype until about age 6.

The development of Lewis antigens is also unique. Lewis antigens are not antithetical, as they result from the interaction of two fucosyltransferases encoded by the Le and Se genes. The Le gene is needed for the production of Lea antigen and the Se gene is needed to form Leb antigen. The three common Lewis phenotypes, Le(a+b-), Le(a-b+) and Le(a-b-) indicate the presence or absence of the Le and Se transferase enzymes.

In pregnancy a mother’s plasma volume increases, and because Lewis antigens are not an integral part of the RBC membrane, they can elute off her RBCs. This causes a decrease in Lewis antigen and some pregnant women, regardless of their true Lewis antigen type, will temporarily type as Le(a-b-). At the same time, because they are now typing Le(a-b-), pregnant women often acquire Lewis antibodies.

Anti-Lea is the most frequently found Lewis antibody, is IgM, and is usually detected at room temperature. In most cases, it is acceptable to give patients with Lewis antibodies RBC units that are crossmatch compatible at 37C without giving antigen negative units. One reason for this is that, as we saw above, Lewis antigens are merely absorbed onto RBCs and can be eluted from transfused red cells within days of transfusion. In addition, when Lewis antigen positive blood is given to Lewis-negative recipients, the Lewis substance in plasma neutralizes antibodies in the recipient. This is why it is extremely rare for anti-Leato cause hemolysis of transfused RBCs. Regardless of Lewis phenotype, RBCs would be expected to have normal in vivo survival.

For an antibody to cause HDFN it must be able to cross the placenta. The antibody must also react with antigens on the red blood cells. Because Lewis antibodies are IgM and do not cross the placenta, and because Lewis antigens are not present on fetal and neonatal erythrocytes, Lewis antibodies have not been implicated in HDFN and this baby is not at risk.

What does this all means in practice? Though the presence of anti-Lewis antibodies in pregnant women is fairly common, both anti-Leaand anti-Leb are naturally occurring IgM antibodies that are not generally considered to be clinically significant. They have low immunogenicity, they do not cause HDFN, they rarely cause hemolysis and do not cause decreased survival of transfused RBCs. This baby is not at risk for HDFN. The mother can safely be transfused with crossmatch compatible RBCs. Her Lea antibodies may be neutralized with a transfusion or will naturally disappear, and her true Lewis phenotype should return within about 6 weeks after delivery.

References

  1. Harmening DM: The Lewis System. In Harmening DM, (6th ed): Modern Blood Banking and Transfusion Practices. FA Davis, Philadelphia 2012, pp. 177-180
  2. Fung, Mark K, ed.: The Lewis System. 18th ed: AABB Technical manual, Bethesda, Md. 2014, pp 304-306
  3. Fung, Mark K, ed.: PreTransfusion testing. 18th ed: AABB Technical manual, Bethesda, Md. 2014, pp 376
  4. D. Radonjic et al, The Presence of antibodies in anti-Lewis system in our pregnant women. Giorn.It.Ost.Gin. Vol. XXXII-n.4.Luglio-Agosto 2010.

 

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: Transfusion Transmitted Malaria

Case Study

A 26 year old African American female with sickle cell anemia presented to a New York emergency room with cough, chest pain, fever and shortness of breath. Laboratory results showed an increased white blood cell count, slightly decreased platelet count and a hemoglobin of 6.2 g/dl. Her reticulocyte count was 7%, considerably below her baseline of 13%. Consulting the patient’s medical records revealed history of stroke as a child and subsequent treatment with chronic blood transfusions. She was admitted to the hospital for acute chest syndrome and aplastic crisis and care was transferred to her hematologist. Two units of RBCs were ordered for transfusion.

The blood bank technologists checked the patient’s blood bank history and noted her blood type was A, Rh(D) positive, with a history of a warm autoantibody and anti-E. The current blood bank sample confirmed the patient was blood type A, RH(D) positive with a negative DAT but the antibody screen was positive. Anti-E was identified. Per request of the hematologist, phenotypically similar units were found and the patient was transfused with 2 units of A RH(negative), C/E/K negative, HgS negative, irradiated blood. The patient’s hemoglobin rose to 8g/dl and she was discharged from the hospital 3 days after transfusion.

Ten days after discharge the patient returned to the emergency room with symptoms including aching muscles, fever and chills. A delayed transfusion reaction was suspected. A type and screen was immediately sent to the blood bank. The post transfusion type and screen remained positive for anti-E, DAT was negative. No additional antibodies were identified. However, a CBC sent to the lab at the same time revealed malarial parasites on the peripheral smear. The patient was consulted for a more complete medical history and reported that she had never traveled outside of the country. A pathology review was ordered and the patient was started on treatment for Plasmodium falciparum.

plasfal1

Discussion

Red Blood cell transfusions can be life saving for patients with sickle cells anemia. These patients are frequently transfused by either simple transfusion of red cell units or by exchange transfusion. Because of this, alloimmunization is reported to occur in 20% to 40% of sickle cell patients.1 Blood bank technologists are very diligent in adhering to strict procedures and follow a standard of practice aimed to prevent transfusion reactions. While preventing immune transfusion reactions may be the most forefront in our minds when transfusing the alloimmunized patient, it is important to consider transfusion transmitted diseases as a potential complication of blood transfusions.

Malaria is caused by a red blood cell parasite of any of the Plasmodium species. Mosquito transmitted infection is transmitted to humans through the bite of an infected mosquito. Transfusion-transmitted malaria is an accidental Plasmodium infection caused by a blood transfusion from a malaria infected donor to a recipient.

Donors, especially those from malarial endemic countries who may have partial immunity, may have very low subclinical levels of Plasmodium in their blood for years. Even these very low levels of parasites are sufficient to transmit malaria to a recipient of a blood donation. Though very rare, transfusion-transmitted malaria remains a serious concern for transfusion recipients. These transfusion-transmitted malaria cases can cause high percent parisitemia because the transfused blood releases malarial parasites directly into the recipient’s blood stream.

Blood is considered a medication in the United States, and, as such, is closely regulated by the FDA. Blood banks test a sample of blood from each donation to identify any potential infectious agents. Blood donations in the US are carefully screened for 8 infectious diseases, but malaria remains one infectious disease for which there is no FDA-approved screening test available. For this reason, screening is accomplished solely by donor questioning.2 A donor is deferred from donating if they have had possible exposure to malaria or have had a malarial infection. Deferral is 12 months after travel to an endemic region, and 3 years after living in an endemic region. In addition, a donor is deferred from donating for 3 years after recovering from malaria. It is important, therefore, for careful screening to take place by questionnaire and in person, to make sure that the potential donor understands and responds appropriately to questions concerning travel and past infection.

Malaria was eliminated from the United States in the early 1950’s. Currently, about 1700 cases of malaria are reported in the US each year, almost all of them in recent travelers to endemic areas. From 1963-2015, there have been 97 cases of accidental transfusion-transmitted malaria reported in the United States. The estimated incidence of transfusion-transmitted malaria is less than 1 case in 1 million units.4 Approximately two thirds of these cases could have been prevented if the implicated donors had been deferred according to the above established guidelines.3 While the risk of catching a virus or any other blood-borne infection from a blood transfusion is very low, a blood supply with zero risk of transmitting infectious disease may be unattainable. With that being said, the blood supply in the United Sates today is the safest it has ever been and continues to become safer as screening tests are added and improved. Careful screening of donors according to the recommended exclusion guidelines remains the best way to prevent transfusion-transmitted malaria.

References

  1. LabQ, Clinical laboratory 2014 No.8, Transfusion Medicine. Jeanne E. Hendrickson, MD, Christopher Tormey, MD, Department of Laboratory Medicine, Yale University School of Medicine
  2. Technical Manual, editor Mark K. Fung-18th edition, AABB. 2014. P 201-202
  3. https://www.cdc.gov/malaria/about/facts.html. Accessed April 2018
  4. The New England Journal of Medicine. Transfusion-Transmitted Malaria in the United States from 1963 through 1999. Mary Mungai, MD, Gary Tegtmeier, Ph.D., Mary Chamberland, M.D., M.P.H., June 28, 2001. Accessed April 2018
  5. Malaria Journal. A systematic review of transfusion-transmitted malaria in non-endemic areas. 2018; 17: 36. Published online 2018 Jan 16. doi: 1186/s12936-018-2181-0. Accessed April 2018
  6. http://www.aabb.org/advocacy/regulatorygovernment/donoreligibility/malaria/Pages/default.aspx

 

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.

Sickle Cell Anemia

Sickle cell disease is a blood cell disorder that affects a lot of Americans, especially those of African descent. The disease is a result of a genetic defect that changes the structure of hemoglobin. This alteration in hemoglobin causes normally round red cells to become sickle in shape as well as sticky and deformed. As a result, these deformed red cells block perfusion of red blood cells into circulation and vital organs and affects the vasculature resulting in severe pain, organ damage and sometimes stroke.

Management of sickle cell disease includes blood transfusion, hydorxyurea and pain management. The long-term administration of hydroxyurea to stimulate the production of fetal hemoglobin is a form of therapy that provides relief in that fetal hemoglobin essentially protects the red cells from sickling. The transfusion of red cells lowers the amount of sickle hemoglobin levels.

Transplantation of hematopoietic stem cells from HLA-identical siblings can be curative in several nonmalignant hematologic disorders, including aplastic anemia, β-thalassemia major, congenital immunodeficiency disorders, and certain inborn errors of metabolism. Pilot studies of bone marrow transplantation for the treatment of young patients with symptomatic sickle cell disease have demonstrated eradication of the underlying disease with low transplantation-related mortality (Bhatia and Walters, 2007).

The process for successful bone marrow transplant to cure sickle cell involves administration of chemotherapy or immunosuppressive drugs to eradicate all the cells.  Now, doctors have developed a more successful regimen where patients take immunosuppressive drugs with a low dose body irradiation, a treatment much less harsh than chemotherapy. Next, donor cells from a healthy and tissue-matched sibling are transfused into the patient. Stem cells from the donor produce healthy new blood cells in the patient, eventually in sufficient quantity to eliminate symptoms. In many cases, sickle cells can no longer be detected. Patients must continue to take immunosuppressant drugs for at least a year.

In the reported trial, published online in the journal Biology of Blood & Marrow Transplantation, physicians from the University in Illinois Chicago transplanted 13 patients, 17 to 40 years of age, with a stem cell preparation from the blood of a tissue-matched sibling. In a further advance of the NIH procedure, the physicians successfully transplanted two patients with cells from siblings who matched but had a different blood type (Parmet, 2015)

Stem cell transplantation has proven itself to provide cures for a lot of hematologic malignancies, now it is successfully finding its way to cure other hematologic abnormalities, including sickle cell anemia.  This continued advancement in medicine will provide relief to a lot of patients who are suffering from sickle cell disease.

References

  1. Bhatia, M., & Walters, M. C. (2007). Hematopoietic cell transplantation for thalassemia and sickle cell disease: past, present and future. Bone Marrow Transplantation, 41(2), 109-117. doi:10.1038/sj.bmt.1705943
  1. Parmet, S. (2015). Adults with sickle cell disease cured with stem cell transplants. Retrieved March 25,2017, from https://news.uic.edu/cure-for-sickle-cell

 

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-Carlo Ledesma, MS, SH(ASCP)CM MT(ASCPi) MT(AMT) is the program director for the Medical Laboratory Technology and Phlebotomy at Rose State College in Midwest City, Oklahoma as well as a technical consultant for Royal Laboratory Services. Carlo has worked in several areas of the laboratory including microbiology and hematology before becoming a laboratory manager and program director.

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.

 

 

Common Thrombotic Microangiopathies

Primary thrombotic microangiopathy (TMA) syndromes encompass diseases that present with thrombosis in small and medium sized blood vessels due to endothelial injury. They are specific disorders that require specific treatment. The initial assessment is focused on confirming that the patient has true microangiopathic hemolytic anemia (MAHA) with or without thrombocytopenia. If MAHA and thrombocytopenia are confirmed it is important to differentiate the primary etiologies, which include:

  1. Thrombotic thrombocytopenic purpura, or TTP, which occurs as a result of severe ADAMTS13 deficiency
  2. Atypical Hemolytic uremic syndrome, or aHUS, which occurs as a result of complement dysregulation
  3. Hemolytic uremic syndrome, or HUS, which occurs as a result of Shiga toxins

TTP results from a severe deficiency of ADAMTS13 (defined as activity <5-10%). ADAMTS13 (a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13) is an enzyme that cleaves large von Willebrand multimers. TTP can be hereditary (called Upshaw-Shulman syndrome) but is usually acquired as a result of an inhibitory autoantibody toward ADAMTS13. The decision to treat TTP is usually clinical since ADAMTS13 assays are often not available and may take several days to return a result if sent to a reference laboratory. TTP typically has more systemic manifestations of organ injury compared to other primary TMA syndromes.

Atypical HUS (aHUS) occurs via a complement-mediated pathway most commonly due to gene mutations of complement factors. Patients with antibodies to complement proteins comprise a smaller proportion of aHUS patients. Penetrance of aHUS is low with only a fraction of gene mutation carriers developing the syndrome. In most cases, a complement activation trigger precedes the manifestation of aHUS.

Most cases of HUS are sporadic, resulting from Shiga toxins produced by Shigella dysenteriae and some serotypes of Escherichia coli (especially O157:H7 and O104:H4). Shiga toxins preferentially injure the renal system by binding to CD77 on kidney epithelial and mesangial cells and endothelial cells. This binding causes downstream ribosomal inactivation leading to programmed cell death (apoptosis). The sporadic form of HUS is associated with bloody diarrhea.

The rationale for focusing on the distinction between HUS, aHUS and TTP is due to the different treatment options for each disease. TTP requires urgent plasma exchange to prevent death, while HUS requires treatment of the infection with little benefit from plasma exchange, while aHUS requires treatment with specialized anti-complement medication. Plasma exchange has inherent risks, so determining the cause of TMA is crucial. While no single clinical feature can be used to determine whether TTP, aHUS, or HUS is responsible for a patient’s symptoms, distinguishing factors include:

  1. Patient age – any primary TMA syndromes may occur at any age; however, children typically present with HUS, aHUS, or hereditary TTP, while adults more commonly present with acquired TTP
  2. Kidney injury –the degree of injury is usually less in TTP than in HUS; aHUS affects the arterioles while HUS tends to affect the glomeruli
  3. Systemic symptoms – up to two-thirds of TTP patients will have some neurologic symptoms; overtly bloody diarrhea is more typical for HUS; aHUS primarily involves kidney injury
  4. A previous episode or family history of aHUS – indication for screening for complement dysregulation in aHUS

 

Want to learn more? Check out: George JN, Nester CM. Syndromes of thrombotic microangiopathy. N Engl J Med. 2014;371(7):654-66.

 

Rogers

-Thomas S. Rogers, DO is a third-year resident at the University of Vermont Medical Center, a clinical instructor at the University of Vermont College of Medicine, and the assistant medical director of the Blood Bank and Transfusion Medicine service.

 

Daratumumab and Blood Bank Testing

Daratumumab, also known as Darzalex, DARA, or Dara-T, is a new medication recently approved in the US by the FDA to treat multiple myeloma. Daratumumab is a novel monoclonal antibody that targets CD38, an integral membrane protein expressed on both plasma cells and red blood cells (RBC). So while the CD38 antibodies are busy destroying the malignant myelomatous plasma cells, they will also be binding onto RBCs. Thankfully, the anti-CD38 binding of RBCs has not been shown to cause severe hemolysis. However, it has been shown to result in false-positive screening test results in the blood bank in all media (saline, PEG, LISS).

The daratumumab effect manifests as a warm autoantibody and will pan-react to any testing carried out including indirect (IAT) and direct antiglobulin tests (DAT), antihuman globulin (AHG) testing, and antibody screening and identification panels. Thankfully, ABO/RhD testing is not affected. To summarize (adopted from AABB Bulletin #16-02):

  1. ABO/RhD typing: no issues.
  2. Immediate spin crossmatch: no issues.
  3. Antibody screen: all cells positive.
  4. Antibody identification panel: all cells positive (autocontrol may be positive or negative).
  5. DAT: positive or negative.
  6. AHG crossmatch: positive with all RBC units tested.
  7. Adsorptions: panreactivity cannot be eliminated.

Potential future techniques to resolve interference, such as anti-idiotype antibodies to neutralize anti-CD38 in vitro, are on the horizon. In the meantime however, our institution’s Blood Bank has asked the clinical teams and pharmacy to notify the Blood Bank if a patient is going to receive daratumumab so that a baseline type and screen and RBC antigen phenotype or genotype is performed prior to initiating treatment. Once the patient receives daratumumab dithiothreitol (DTT) is used to eliminate the CD38 antigen from the surface of the reagent RBCs thereby eliminating the antibody screen and panel panreactivity. DTT treatment also destroys antigens in the Kell family. Therefore, unless the patient is known to be Kell-positive by phenotype/genotype, Kell-negative units are provided to patients on daratumumab.

Communication between clinicians, pharmacy and Blood Bank when a patient is receiving daratumumab is paramount to prevent delays in Blood Bank testing and transfusion needs. Protocols to handle Blood Bank specimens from patients receiving daratumumab can help streamline testing and reduce turnaround time for transfusion.

For further reading check out the AABB Bulletin #16-02 issued this year.

 

Rogers

-Thomas S. Rogers, DO is a third-year resident at the University of Vermont Medical Center, a clinical instructor at the University of Vermont College of Medicine, and the assistant medical director of the Blood Bank and Transfusion Medicine service.

FDA Issues Revised Recommendations for Reducing the Risk of Zika Virus Transmission through Transfusion

Today, the FDA released industry guidance for reducing the risk of Zika Virus transmission through blood products. “Revised Recommendations for Reducing the Risk of Zika Virus Transmission by Blood and Blood Components”  is for immediate implementation.