Transfusion Associated Graft-Versus-Host Disease

Transfusion-associated graft-versus-host disease (TA-GVHD) is a rare complication that develops 4 to 30 days after the transfusion of cellular blood products (i.e. red blood cells, platelets, granulocytes). It can occur in both immunocompromised and immunocompetent patients, and recognition is often delayed because the nonspecific symptoms are attributed to the patient’s underlying diagnosis. TA-GVHD affects the transfusion recipient’s bone marrow and is nearly universally fatal, making prevention absolutely essential.

TA-GVHD is mediated by viable mature immunocompetent donor lymphocytes against the recipient’s antigen presenting cells. TA-GVHD does not occur after most transfusions because the donor lymphocytes are destroyed by the recipient’s immune system before they can mount a response against the host. However, this protective response does not occur in certain settings. One is profound cell-mediated (T cell) immune deficiency, resulting from congenital, acquired, or iatrogenic causes. Another occurs when there is a specific type of partial Human Leukocyte Antigen (HLA) matching between the donor and recipient. HLA molecules are the primary means of distinction between self and non-self. If the donor HLA phenotype is homozygous and the recipient expresses the same HLA haplotype, it may mask donor lymphocytes from the recipient. The end result is engraftment and proliferation of mature donor T cells in the recipient’s bone marrow.

The donor T cells are then activated by mismatched HLA class I major antigens. This immunologic assault typically manifests clinically with fever and an erythematous, maculopapular rash which often progresses to generalized erythroderma. In addition to skin dysfunction, liver, gastrointestinal tract, and bone marrow symptoms are also common. The main laboratory findings of TA-GVHD include pancytopenia due to hypocellular marrow, abnormal liver function tests, and electrolyte abnormalities induced by diarrhea.

The differential diagnosis of TA-GVHD is broad. A more definitive diagnosis is suggested from skin biopsy which classically reveals vacuolization of the basal layer and a histiocytic infiltrate, and occasionally shows an almost pathognomonic finding — satellite dyskeratosis, which is characterized by single, dyskeratotic cells accompanied by lymphocytes. The definitive diagnosis of TA-GVHD relies in demonstrating that circulating lymphocytes have a different HLA phenotype from recipient APCs, proving that they came from the donor.

As mentioned above, TA-GVHD portends a high mortality rate and is poorly responsive to the available therapies; therefore prevention is of primary importance. Current strategies include gamma irradiation or leukocyte inactivation (i.e. pathogen reduction technology) of the blood products prior to transfusion to disable donor lymphocytes. Some of the more common indications for patients requiring irradiated blood products include those who are immunosuppressed, who have received a hematopoietic cell transplant, who are receiving blood components from a related donor, or who are given HLA-matched platelets. There is also evidence that transfusing older products decreases the risk of TA-GVHD due to the shortened lifespan of T cells within the products. In summary, TA-GVHD can occur in both immunocompetent and immunocompromised recipients, is mediated by donor T lymphocytes, and is almost always fatal.

For further reading, please see the review article by Kopolovic et al. A systematic review of transfusion-associated graft-versus-host disease. Blood. 2015;126(3):406-14.



-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 Halts Blood Donation in Two Florida Counties Due to Zika Virus

From the Washington Post:

“In a notice sent to blood centers and posted on the agency’s website Wednesday evening, the FDA said it is requesting all blood centers in Miami-Dade and Broward counties to ‘cease collecting blood immediately’ until those facilities can test individual units of blood donated in those two counties with a special investigational donor screening test for Zika virus or until the establishments implement the use of an approved or investigational pathogen-inactivation technology.”


A Brief Overview of 7-day Platelets

The transfusion community has targeted platelets as the primary culprit in transfusion-associated clinical sepsis and fatal microbial infection. Platelets (PLTs) are associated with a higher risk of sepsis and related fatality than any other transfusable blood component. Concerns over bacterial contamination in PLT concentrates prompted the US Food and Drug Administration (FDA) in 1986 to issue a memorandum limiting the storage time of platelet products to 5 days. Only recently did the FDA issue draft guidance describing bacterial testing to improve the safety and availability of PLTs, and outlined the steps necessary for transfusion services to extend apheresis PLTs to 7 days.

Microbial infections were the 4th leading cause of transfusion-related mortality, accounting for 8% of them between 2010 and 2014. PLT storage at ambient room temperature supports high titer bacterial proliferation. Skin flora are the most common source of contamination, occurring at the time of collection. Despite the introduction of improved pre-collection arm preparation and analytically sensitive culture-based bacterial detection methods, the risk of fatal and non-fatal clinical sepsis has persisted.

Most recently, the 2016 AABB standards stated that PLTs may be stored for 7 days only if: 1) storage containers are cleared or approved by FDA for 7-day PLT storage and 2) labeled with the requirement to test every product stored beyond 5 days with a bacteria detection device cleared by FDA and labeled as a “safety measure.” The Verax PGD test is a rapid, single use, lateral flow immunoassay, and the only rapid, day of transfusion test the FDA has cleared as a “safety measure.” The proprietary test detects surface bacterial antigens, namely lipotechoic acid found on gram positive organisms and lipopolysaccharide found on gram negatives. The PGD test as a “safety measure” is to be used in concert with culture, not replace it.

Verax PGD test

Approximately 2.2 million PLT transfusions are administered yearly in the United States, of which more than 90% consist of apheresis PLTs. If the available data were generalized to the entire US apheresis PLT supply, approximately 650 contaminated apheresis PLTs would be caught with the PGD test, preventing septic transfusion reactions and potential fatalities each year. The FDA approval of this test allows non-culture based testing to extend dating from 5 to 7 days and further closes the safety gap that exists in apheresis PLTs.



-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.

The author declares that he has no disclosures.

Cryoprecipitate 101

Cryoprecipitate, or cryo for short, is a fresh frozen plasma (FFP)-derived concentrate including fibrinogen, factors VIII and XIII, von Willebrand factor, and fibronectin. Cryo contains only 40-50% of the coagulation factors found in a unit of plasma but is concentrated into a reduced 15-20 ml volume. Cryo is prepared from FFP as it is thawed slowly at 4° C. A precipitate forms at the bottom of the bag, which is then separated from the supernatant plasma. Cryo is stored frozen at at least 18° C and must be transfused within 6 hours of thawing or 4 hours of pooling. Each unit from a separate donor is suspended in 15 mL plasma prior to pooling.

Dose per unit



150-250 mg

100-150 hours

Von Willebrand factor

100-150 U

24 hours

Factor VIII

80-150 U

12 hours

Factor XIII

50-75 U

150-300 hours

Cryo is used most commonly for replacement of fibrinogen in patients that are bleeding or at increased risk of bleeding. Fibrinogen replacement may be indicated for hypofibrinogenemia (fibrinogen < 100 mg/dL) or dysfibrinogenemia. The target increase in fibrinogen level is 30-60 mg/dL in adults and 60-100 mg/dL in pediatric patients. Many institutions transfuse cryo prior to administration of factor VIIa concentrate to ensure adequate fibrinogen for clot formation given the cost and short half-life of factor VIIa of about 4 hours. Fibrinogen replacement can be monitored with a fibrinogen level assay and clinical response.

Cryo may be used to treat von Willebrand disease, Hemophilia A (factor VIII deficiency), or Factor XIII deficiency only when the appropriate plasma-derived or recombinant factor concentrates are unavailable and/or desmopressin (DDAVP) is ineffective or contraindicated. Cryo is sometimes useful if platelet dysfunction associated with renal failure does not respond to dialysis or DDAVP. Cryo also contains fibronectin; however there are no clear indications for fibronectin replacement.

Topical application of cryo in combination with thrombin as a “fibrin glue” has been used as a surgical hemostatic agent. This application is being discontinued due to the preferred commercially available virus-inactivated fibrin sealants with higher fibrinogen concentrations.

Historically, the dosing was a 10-unit pool for adults and 1-2 units/10kg for pediatric patients based on fibrinogen content. However, Blood Bank and Transfusion services should check with their blood supplier on actual fibrinogen content in individual and pre-pooled units as the fibrinogen content has likely increased (~325 mg) due to improved preparation. Therefore Blood Bank and Transfusion services can probably decrease the standard dose to 4-5 pooled units for adults and 1 unit/10 kg for kids.

A previous version of this post said that cryo is frozen at 1-6°C; this is incorrect. The correct temperature is 18°C, and has been corrected in the text. Thank you, astute readers, for correcting our errors! –Lablogatory editors



-Thomas S. Rogers, DO is a second-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.

Collaboration is King

In the April issue of Transfusion journal, Joseph et al report their 1½ -year experience with the use of 4 Factor Prothrombin Complex Concentrate (4F-PCC) for urgent reversal of Vitamin K antagonists (Transfus 2016;l 56: 799-807).

As the authors mention, their “…study supports the safety of 4F-PCC for urgent vitamin K antagonist reversal even in unselected patients.”

I highlight this article for several reasons. It is incumbent upon those of us in the clinical laboratory, and especially the Blood Bank/Transfusion Service, to be aware of these new pharmaceutical agents that help provide rapid reversal of anticoagulants and allow for the potential elimination of unnecessary transfusions. I have found that often our clinical colleagues are unfamiliar with these strategies and we must take the lead in helping to establish protocols for their appropriate use. This article speaks, as well, to the need for ongoing evaluation of these drugs in, as they state in their title, the “real-world” of medical practice. Knowing how specific drugs affect outcomes outside of select studies with exclusions of particular patient populations (in this case, those with TE risk) is so valuable to our everyday work.

Another reason that this article is important is it underscores the importance of collaboration. The authors are representatives of departments of Pathology, the School of Medicine and Pharmacy. It is vital that we, as laboratory professionals, push to participate alongside our clinical colleagues in the assessment and implementation of new therapies and adjuvant treatments.

It is obvious from the Transfusion Medicine perspective, that our Pharmacy “friends” play a huge role in patient care, often spearheading and specializing in areas such as anticoagulant reversal strategies, release of factor concentrates, antifibrinolytics, IVIg and albumin. All of these pharmaceuticals can ultimately affect our laboratory testing and our potential interventions. Be certain you have representative from Pharmacy as a member of your Transfusion Committee.

It always pleases me to see, not only excellent literature, but also ongoing collaboration with laboratory professional often at the helm!



-Dr. Burns was a private practice pathologist, and Medical Director for the Jewish Hospital Healthcare System in Louisville, KY. for 20 years. She has practiced both surgical and clinical pathology and has been an Assistant Clinical Professor at the University of Louisville. She is currently available for consulting in Patient Blood Management and Transfusion Medicine. You can reach her at

How We Treat Weak D and Partial D Transfusion Recipients

The weak D (formerly Du) phenotype describes an individual with a variant RHD allele, leading to low expression of complete D antigen on the surface of their red blood cells. The partial D phenotype describes a variant RHD allele that results in modification of the surface D antigen and can result in the loss of D epitope. The prevalence of these phenotypes is thought to occur in 0.2 to 1% of Caucasians. Typically, alloimmunization with anti-D is more likely to occur in partial D individuals who are exposed to the Rh(D) antigen than in weak D individuals. However, there are instances when weak D individuals may develop an anti-D alloantibody after exposure to Rh(D) positive blood.  Since the most common methods of immunohematology testing in the Blood Bank cannot reliably discern between weak D and partial D expression, a standard practice is treat both weak D and partial D individuals as Rh(D) negative when they are recipients of blood products.

The AABB Standards for Blood Banks and Transfusion Services does not require that weak D testing be performed on Rh(D)-negative recipients of blood products. In line with this, our transfusion service does not routinely perform serologic weak D testing on transfusion recipients and our testing algorithm is designed to consider weak D and partial D individuals as Rh(D) negative for transfusion purposes. We believe our testing strategy helps prevent anti-D alloimmunization in a vulnerable population, especially women of childbearing potential, and helps streamline test utilization in the Blood Bank. That said, we continue to perform weak D testing on potential red blood cell donors (i.e. fetus and newborn of Rh(D) negative mother, stem cell and solid organ donors).

Rh typing discrepancies may occur in the following situations:

  1. Obstetric patients: A patient typed as weak Rh(D) positive during her prior pregnancy and did not receive Rh immune globulin prophylaxis (RhIg). However, during the current pregnancy, this patient is now typed as Rh(D) negative due to our updated procedure. If the newborn is Rh(D) positive, a fetal screen and Kleihauer-Betke test will be performed as needed, and an appropriate RhIg dose is recommended.
  2. Previous blood donors, organ donors, and cord blood from neonates: It is important to identify the weak D phenotype in blood donors (including cord blood from neonates) since very low levels of D antigen are sufficient to elicit the formation of an anti-D alloantibody in Rh(D) negative transfusion recipients. A patient who was previously a blood donor would be typed by the blood collection center as Rh(D) positive due to the presence of weak D, but as a transfusion recipient would be typed as Rh(D) negative since weak D testing is not performed on transfusion recipients.



-Thomas S. Rogers, DO is a second-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.

A Strategy for Patients with Sickle Cell Disease

Transfusion of red blood cells (RBCs) is a cornerstone of treatment to prevent the complications of sickle cell disease (SCD). SCD is caused by a mutation of the β-globin gene, resulting in glutamic acid being substituted by valine at position 6. The mutation results in an abnormal hemoglobin (Hb SS) that aggregates into a rigid sickle-shape under certain conditions. Individuals with SCD frequently require transfusion of RBCs to treat acute pain crisis (i.e. acute chest syndrome) and prevent chronic complications (i.e. stroke). RBC transfusion helps SCD patients by providing RBCs with hemoglobin A thus decreasing the amount of HbSS RBCs that can sickle and contribute to pain crisis and chronic complications. Unfortunately, alloimmunization to non-ABO RBC antigens is a potential complication of any patient receiving chronic transfusion therapy. The most life-threatening consequence of alloimmunization in SCD is the development of a delayed hemolytic transfusion reaction with hyperhemolysis. Alloimmunization also puts SCD patients at increased risk of receiving an incompatible transfusion due to difficulty in finding compatible blood and increases costs for the health care system.

Antigen matching beyond standard ABO and Rh typing can help reduce the alloimmunization rate in chronically transfused patients. A widely accepted antigen matching strategy used by transfusion services is to initially provide Rh and Kell-matched RBC units to SCD patients, even if the patient has not yet made an alloantibody (i.e. antibody screen negative) since the Rh (D, C, c, E and e) and Kell (K) antigens are among the most immunogenic.  Providing Rh and K-matched RBC units continues until the patient proves to be an antibody former (i.e. anti-Jk(b)), after which the transfusion service provides fully phenotype matched RBCs for non-emergent transfusion when available.  A “full” phenotype usually includes Rh, K, Jk(a), Jk(b), Fy(a), Fy(b), M, N, S and s.

In summary, the strategy for patients with SCD is as follows:

  1. Determine the patient’s full RBC phenotype (D, C, c, E, e, K, Kidd, Duffy, M, N, S, s) before transfusions begin. If transfusions already started, consider molecular testing.
  2. Provide Rh (D, C, c, E, e) and K-matched RBCs until the patient proves to be an antibody former.
  3. If the patient proves to be an antibody former, provide full phenotype matched RBC units to attempt to prevent any additional antibody formation and it becomes increasingly impossible to find compatible units.



-Thomas S. Rogers, DO is a second-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.