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An update on blood typing, crossmatching, and doing no harm in transfusing dogs and cats
Seven blood types are recognized in dogs, and four blood types are identified in cats.
Blood types are classifications of heritable species-specific antigens on the surface of red blood cells. Seven blood types are recognized in dogs, and four blood types are identified in cats. Other cells such as leukocytes, platelets, or cells in other tissues may also share these antigens. Alloantibodies (or isoantibodies) are antibodies present in the serum against an antigen from another animal of the same species. These may be naturally acquired (e.g. ingestion of colostrum) or induced through previous exposure (e.g. transfusion), and their presence is detected by a crossmatch.
Blood product transfusion may produce a wide range of harmful effects in veterinary patients. Some of these effects are common and may be unavoidable (e.g. fever), but others, such as immune-mediated acute and delayed transfusion reactions that are directly associated with inappropriate type and crossmatch processes in dogs and cats, can be minimized.
In this article, I present an overview of blood typing in dogs and cats and proper crossmatching techniques. I also offer decision-making recommendations for veterinarians to help avoid transfusion reactions, and I discuss the signs that may be observed if a reaction occurs.
CANINE BLOOD TYPES AND ANTIBODIES
Dog blood types are numbered according to the dog erythrocyte antigen (DEA) system.
DEA 1.1, 1.2, and 1.3
DEA 1 was formerly known as A and consists of four alleles: negative, 1.1, 1.2, and 1.3. DEA 1.1 is inherited as an autosomal dominant trait over DEA 1.2, and the null type is recessive to both. DEA 1.1 and DEA 1.2 are the most important antigens and together occur in about 60% of dogs.1 Confusion may arise because both of these types have been considered A positive; however, DEA 1.2 dogs, which make up 7% to 29% of dogs, will develop potent anti-DEA 1.1 antibodies once transfused with DEA 1.1 cells.
While naturally occurring antibodies to these antigens are generally considered nonexistent, first-time transfusions with DEA 1.1 blood may be associated with a decreased circulating lifespan of transfused cells, and subsequent transfusions will be associated with an acute hemolytic reaction. Transfusion of DEA 1.2 blood to a sensitized DEA negative dog will result in an exponential loss of cells over the course of several weeks, with about half of the transfused cells being lost within the first 10 days.2 DEA 1.3 is only known to exist in dogs from Australia, primarily German shepherds.3
DEA 4 occurs in up to 98% of dogs, and dogs with this type alone are considered universal donors. Only about 75% of Doberman pinschers are DEA 4 positive. Naturally occurring DEA 4 antibodies are not known to exist; however, hemolytic transfusion reactions can occur after sensitization with DEA 4 positive blood transfusions in dogs lacking that antigen.4
DEA 3 and 5
DEA 3 and 5 are expressed in lesser proportions of the dog population, but DEA 3 occurs in 23% of greyhounds, and 30% of greyhounds are DEA 5 positive. Naturally occurring antibody is present in 20% of DEA 3 negative dogs and 10% of DEA 5 negative dogs in the United States.2
DEA 7 is present in 8% to 45% of U.S. dogs. Naturally occurring antibodies have been observed against DEA 7, with a delayed transfusion reaction causing the decreased lifespan of transfused cells but no hemolysis.5,6 While controversy regarding the importance of the antigen exists, it is best to avoid the premature loss of transfused cells by using donor blood lacking this antigen.
A new antigen was reported in 2007 and found to be present in about 93% of U.S. dogs.7 It was temporarily named Dal because the index case involved a Dalmatian. The Dalmatian was typed as DEA 1.1, 3, 4, and 5 positive and DEA 7 negative but became sensitized after multiple transfusions for chronic renal failure with blood typed as DEA 1.1, 4 positive only. Because additional transfusions were necessary, compatibility testing was required. Major crossmatches between the index dog and 55 non-Dalmatian donors that should have been compatible based on types DEA 1.1, 1.2, 3, 4, 5, and 7 were incompatible. Major crossmatches between the index dog and only 20 of 25 (80%) unrelated Dalmatians were compatible. Incompatible transfusions involving this antigen could result in acute and delayed hemolytic reactions. When transfusions become necessary for sensitized Dalmatians, it appears that compatible donors will most likely be found within the Dalmatian breed.
Little is known about DEA 6 and 8 and about 11 other antigens thought to exist because typing sera for these antigens are not available. Without typing sera for these antigens, their relationship to Dal could not be determined.
FELINE BLOOD TYPES AND ANTIBODIES
In cats, only the AB system has been routinely recognized and consists of three types: A, B, and AB.
Type A is the most common and occurs in more than 95% of domestic shorthaired and longhaired cats in the United States. To date, all Siamese, Burmese, Tonkinese, Russian blue, American shorthair, and Oriental shorthair cats have been identified as type A.8-10 Type B has been identified in up to 10% of Maine coon and Norwegian forest cats; up to 20% of Abyssinian, Birman, Persian, Somali, Sphinx, and Scottish fold cats; and up to 45% of exotic and British shorthair, Cornish rex, and Devon rex cats. Type AB has been observed in domestic shorthaired cats as well as in breeds with type B.11
This blood system follows simple Mendelian inheritance with the A (A) gene having dominance over the AB (ab) gene, which has dominance over the B (b) gene. Type A cats may have any one of three genotypes: A-A, A-ab, or A-b. Type AB cats may have either an ab-ab or ab-b genotype, and a type B cat can only have the b-b genotype. Thus, a breeding pair of type A cats can produce kittens of types A, AB, or B, depending on their phenotypes.
Unlike dogs, cats have marked naturally occurring antibodies. All type B kittens develop antibodies within a few weeks after birth, and high titers develop by three months of age.12 As a result, type B queens will have strong anti-A antibodies in their colostrum without any prior exposure from pregnancy or transfusion. Type A kittens will also develop antibodies, but these are generally considered less potent. Since antibodies can be transferred to a kitten through the colostrum for up to 16 hours after birth, kittens born healthy can suddenly fade from the hemolytic anemia that develops. This hemolytic anemia generally occurs in type A or AB kittens born to B queens mated with type A toms.13
Type AB cats are considered to be universal recipients since they lack anti-A and anti-B antibodies; however, they should be transfused with type A cells to avoid inadvertently transfusing potent anti-A antibodies from a type B donor, which is an example of a minor side reaction. Because of the effects of geography and breed on the frequency of blood types, the risk of inducing a potentially fatal transfusion reaction in type B recipients could be as high as 20% when transfusing unmatched blood.
A new antigen, Mik, was reported in 2007 and is present in many domestic shorthaired cats.14 Cats lacking this antigen (about 6% of those tested) have the potential to develop an acute hemolytic reaction after the transfusion of AB matched blood. Since typing serum is not available for the Mik antigen and antibodies appear to be naturally occurring, crossmatching even type-matched cats before a transfusion is prudent.
BLOOD TYPING AND CROSSMATCHING
Freshly collected blood in EDTA and a clot or plain tube from both the recipient and donor are recommended for typing and crossmatching unless the donor has been screened for antibodies before, in which case only donor cells from the EDTA sample are needed. Alternatively, tubing segments ("pigtails") (Figure 1) may be used from the donor unit as long as the unit's sterility remains intact. The samples should be free of hemolysis and lipemia.
Figure 1. "Pigtails" from the donor unit may be used to blood type and for crossmatching with the recipient as long as the unitâs sterility remains intact. (Photo by Charlie Kerlee.)
Commercial blood typing kits are available for dogs and cats and can be used to screen potential donors and to make appropriate selections for crossmatches and transfusions based on the recipient's blood type. Examples include typing cards (DMS Laboratories) and an immunochromatographic cartridge (Alvedia) (Table 1). These kits type for DEA 1.1 only in dogs and for A, B, and AB in cats. Both the cards and cartridge are relatively simple typing methods, requiring only a few minutes to run, and include a means of performing an auto control to identify potential interference from autoagglutination. Using a 2+ agglutination endpoint reaction for the cards, one study obtained three false negative and five false positive reactions out of 88 dog samples tested.15 This issue has reportedly been corrected.16 In the same study, the cartridge obtained no false negative and six false positive results. A gel column diffusion assay test (DiaMed) used in that study is no longer available for the veterinary market.
Table 1: Selected Blood Typing and Blood Product Websites
Erroneous results can be obtained with failure to follow kit instructions. Autoagglutination and cross contamination from previously used stir sticks can cause false positive results with card typing methods. False negative results can be obtained with blood from extremely anemic animals (PCV < 10%) and from a prozone reaction (excess antibody for amount of antigen present).17
A recent study suggests a more thorough, extended typing kit may become available that will type for DEA 1.1, 3, 4, and 7 and Dal.18 An appropriate dilution for the DEA 1.2 typing reagent was not identified, and DEA 5 was not included. In this study, 10 dogs received DEA 1.1 compatible transfusions and were crossmatched before and after transfusions. Six of the crossmatch pairings in four of the dogs could have developed antibodies based on the typing results, and four crossmatches involving two dogs became incompatible 21 to 23 days later with reaction strengths ranging from 3+ to 4+. A third dog had a 1+ incompatibility at day 13 that became compatible by day 50. Five crossmatch pairings in four dogs were not expected to develop antibodies based on the extended typing results; however, major incompatible crossmatch results were obtained ranging in strength from 1+ to 3+ over the course of two to four weeks. These incompatible results indicate sensitization to antigens not detected by the typing process (e.g. DEA 5, 6, 8).
Even though these typing methods are relatively simple, read the package inserts thoroughly for sources of potentially erroneous results and follow the instructions exactly. When confirmatory testing is necessary, such as for selecting permanent donors, for checking questionable results, or in lieu of in-house typing for elective surgeries, outside laboratories such as Animal Blood Resources International in Stockbridge, Mich., or the University of Pennsylvania's Hematology and Transfusion Laboratory may be used. The extended blood typing kit option may expand these possibilities to other testing sites.
A major crossmatch tests for detectible naturally occurring or induced antibodies in the recipient serum against donor erythrocytes. This testing should be done any time a patient is likely to have relevant naturally occurring antibodies (cats), if the patient's transfusion history is unknown, or if a transfusion occurred at least two to four days previously, even if it was with the same donor.1,8,19,20 Commercial crossmatch kits are available from DMS Laboratories.
A minor crossmatch tests if detectible antibodies are present in the donor plasma or serum against the patient erythrocytes. While considered less important, minor side reactions occasionally occur. Permanent donors can be selected based on commercially offered blood typing reagents and screening for antibodies in order to minimize the chance of a minor side reaction. The typing and crossmatch kits typically provide controls to rule out false positive reactions because of autoagglutination or claim no interference from it.
A slide crossmatch is a crude method of crossmatching that should be reserved for emergency situations only. In this case, the major slide crossmatch consists of mixing two drops of recipient plasma with a drop of blood from the donor at room temperature on a clean glass slide and observing for agglutination while rotating the slide for one minute. A minor crossmatch can be performed in the same way using two drops of donor plasma and one drop of recipient blood. However, two potentially serious errors can occur with this method. First, potentially fatal hemolytic reactions may be missed since hemolysis is difficult to recognize by using this method. Second, this procedure can miss prozone reactions where the presence of excess antibody for the amount of antigen can result in failure of agglutination.
DETECTING A TRANSFUSION REACTION
An acute intravascular hemolytic transfusion reaction can occur in type B cats receiving type A blood. A severe reaction most commonly occurs in dogs previously sensitized to DEA 1.1 blood, but it has also been reported in dogs sensitized against DEA 4, Dal, or an unidentified type (non-DEA 1.1, 1.2, 3, 4, 5, 7).1,4,5,7 Signs associated with acute hemolytic transfusion reactions typically begin immediately after initiating the transfusion and may include fever, altered heart rate, hypotension, dyspnea, loss of bladder and bowel control, vomiting, hemoglobinemia, and hemoglobinuria.21 Since the transfused cells undergo hemolysis, the packed cell volume (PCV) fails to rise. Sequelae can include disseminated intravascular coagulation, renal failure, shock, and death. The reaction's severity is associated with the antibody titer and the amount of blood transfused. If any of the signs beyond fever are noted, the transfusion should be stopped and the appropriate therapy should be initiated.
The half-life of type-compatible blood administered to dogs and cats is about three and five weeks, respectively. Delayed transfusion reactions are more insidious than acute reactions and may be overlooked or attributed to other events, such as an antibiotic-related allergy. In these cases, the PCV may rise as expected and then fall over the course of several days to weeks. Since the hemolysis is extravascular, icterus and hyperbilirubinuria may be noted.
A first-time type B transfusion to a type A cat may result in a delayed hemolytic reaction. Delayed reactions may also occur in first-time canine recipients of DEA 1.1 or DEA 7 blood that are negative for those antigens as well as in dogs previously sensitized against weaker antigens. Donors deemed DEA 1 negative may actually be type DEA 1.2 if one relies solely on the in-house typing kit results. Ideally, permanent donors should receive complete typing and antibody screening if possible. The in-house DEA 1.1 typing kits should be reserved for screening potential donors and typing recipients requiring immediate transfusion therapy.
A dog receiving blood type-matched for DEA 1.1 can still be mismatched for any of the other antigens. Universal donors should be positive for DEA 4 only since it is a common antigen and will induce sensitization only in the rare dog that lacks it. However, it is important to remember that universal donor blood is only known to be negative for DEA 1.1, 1.2, 3, 5, and 7. The elusive DEA 6 and 8 and several other antigens for which typing sera do not exist can be present on universal donor erythrocytes and can sensitize a recipient negative for any one or more of those antigens. The Dal antigen also needs to be kept in mind when transfusing Dalmatians.
GUIDE FOR TYPING AND CROSSMATCHING DOGS
Crossmatching may not detect low concentrations of antibody or predict the potential to develop antibodies against mismatched blood. Repeat transfusions may be a rare occurrence in some practices, but for those with a high volume of patients with chronic renal disease or receiving chemotherapy or those with tenacious owners battling persistent immune-mediated hemolytic anemia in their pets, repeat transfusions are not uncommon. In these cases in which multiple transfusions are anticipated, donor selection based on blood type can maximize transfusion effectiveness.
Table 2: Process for Typing and Crossmatching Dogs
Following a few easy steps (Table 2) can help you decide whether a simple DEA 1.1 typing is sufficient or if a more complete typing is needed and whether crossmatching dogs to minimize transfusion reactions and sensitization to transfused blood is necessary. Briefly, DEA 1.1 type-matched blood is adequate for a first-time recipient requiring immediate transfusion therapy. Even if sensitization occurs, the benefits of the transfusion likely outweigh the shortened half-life of transfused cells. Universal donor blood is recommended as a first choice whenever possible and, certainly, when repeated transfusions are anticipated. Crossmatching will not be helpful in a first-time recipient (or recent recipient) but is required if the recipient has received a transfusion four or more days previously or is a Dalmatian. Common antigens that are lacking in a sensitized recipient can make locating compatible donor blood challenging. In these cases, crossmatching against siblings or same-breed donors is more likely to be successful.
Linda M. Vap, DVM, DACVP
Department of Microbiology, Immunology and Pathology
College of Veterinary Medicine and Biomedical Sciences
Colorado State University
Fort Collins, CO 80523
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