Articles on Medical Diseases and Conditions

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  • Anaphylactic Reactions

    Immunoglobulin A antigen reactions. Immunoglobulin A (Chapter 22)is the principal immunoglobulin in such human secretions as saliva, bile, and gastric juice. Class-specific anti-IgA occurs in patients who lack IgA; these persons may be clinically normal or may have such disorders as malabsorption syndrome, autoimmune disease, or recurrent sinus or pulmonary infection. Interestingly, only 40% develop anti-A antibodies. Limited specificity anti-IgA occurs in persons who have normal IgA levels but who become sensitized from exposure to human plasma proteins from blood transfusion or pregnancy. Anti-IgA antibodies produce reaction only in transfusions that include human plasma proteins and are considered to be a type of anaphylactic reaction. Symptoms consist of tachycardia, flush, headache, dyspnea, and sometimes chest pain. Typically there is no fever. Severe episodes may include hypotension. Again, these are nonspecific symptoms that could be produced by leukoagglutinins or hemolytic reactions. Tests to prove IgA incompatibility are available only in a few medical centers; therefore, the diagnosis is rarely confirmed. Substantially decreased IgA on serum IgA assay would be presumptive evidence of anti-IgA reaction of the class-specific type. IgA immunologic reactions can be prevented by eliminating donor plasma proteins through the use of washed cells or frozen cells.

  • Tissue-Organ Immunologic Reactions

    Graft-vs.-host disease (GVHD)

    Graft-vs.-host disease (GVHD) results from introduction of sufficient HLA-incompatible and immunologically competent donor lymphocytes into a recipient who is sufficiently immunodeficient that the incompatible donor cells cannot be destroyed. The donor lymphocytes proliferate and attack tissues of the new but incompatible host. There are two clinical types, organ transplants (represented here by bone marrow transplant) and blood transfusion. In bone marrow grafts the donor lymphocytes are in the graft. Current reports indicate that 60%-70% of patients with bone marrow grafts from HLA-compatible siblings (not identical twins) develop some degree of GVHD symptoms, and 10%-20% of marrow transplant patients die from it.

    Graft-vs.-host(GVH) transplant disease can be acute or chronic. In bone marrow transplants, the acute type clinically begins about 10-28 days after transplantation; the first symptom usually is a skin rash. This is a somewhat longer time interval than onset in transfusion-related GVHD. About 20%-50% of HLA-compatible marrow transplant patients develop some degree of acute onset GVHD. Chronic GVHD occurs in 25%-45% of longer-term marrow transplant survivors; it typically appears 100 days or more after transplantation, but it can occur as early as 50 days or as late as 15 months. Persons under age 20 years have a relatively good chance of escaping or coping with GVHD, while those over age 50 have the worst prognosis. Marrow transplant patients develop B-lymphocyte lymphoproliferative disorders (benign, premalignant, or malignant lymphoma) in about 0.6% of cases. This is often associated with evidence of Epstein-Barr viral infection.

    In the other clinical category of patients, transfusion is the usual cause. Patients at risk are those receiving intrauterine or neonatal exchange transfusions, and in congenital immunodeficiency syndromes, malignancy undergoing intensive chemotherapy (especially when combined with radiotherapy), and aplastic anemia. Incidence of posttransfusion graft-vs.-host syndrome in these conditions is not known with certainty but varies from 0%-8%, depending on the disease, therapy, and institution. The highest incidence appears to be in malignant lymphoma. In classic cases the syndrome develops 1-2 weeks (range 3 days-6 weeks) after transfusion and consists of fever, severe diarrhea, pancytopenia, hepatitis, and skin rash. Up to 90% of patients with a transfusion-related full-blown syndrome die, usually from infection. Both transplant and transfusion syndromes can be prevented by gamma radiation treatment of the blood product being transfused, in doses sufficient to affect lymphocytes but not other blood cells (usually 25 Gy = 2,500 rads). The AABB recommends radiation for all donors related by blood to the recipient. The greatest incidence of GVHD from blood-related donors is from first-degree blood relatives. RBC transfusion itself has immunologic effects on the recipient. It has been shown that random-donor blood transfusions before renal transplant decrease the incidence of graft rejection. Some institutions use RBC transfusion for this purpose. It is reported that transfusion therapy depresses natural killer cell function in these patients but the T4/T8 cell ratio remains normal. In addition, some studies report a somewhat greater incidence of recurrences or decreased overall survival in colon cancer patients who had multiple transfusions compared with those without transfusion, although this is disputed by other investigators.

  • Cytopenic Reactions

    Thrombocytopenia

    Blood platelets contain at least three antigen systems capable of producing a transfusion reaction. The first is the ABO (ABH) system, also found on RBCs; for that reason the AABB recommends that single-donor platelet units be typed for the ABO group before being transfused. Platelets do not contain the Rh antigen; however, since platelet units may be contaminated by RBCs, it is safer to type for D (Rho) antigen in addition to ABO typing. The majority of investigators believe that ABO-Rh typing is useful only to avoid possible sensitization of the recipient to RBCs and that recipient anti-A or anti-B antibody will not destroy ABO-incompatible platelets. There are platelet-specific antigens (PLA1 group, also called Zwa, and several others) that are found in high incidence and thus rarely cause difficulty. However, PLA1 antibodies have been implicated in a syndrome known as posttransfusion purpura (Chapter 8). Finally, there is the class I HLA-A,B,C group. This tissue compatibility system has been incriminated in patients who become refractory to repeated platelet transfusions (i.e., in whom the platelet count fails to rise) and in those who develop a febrile reaction after platelet administration. Interestingly, it has been reported that there is not a good correlation between the number of platelet transfusions and development of HLA antibodies. Persons who are HLA-compatible (usually siblings) as a rule are able to donate satisfactorily. The HLA-A antigens are also present on WBCs. Finally, other types of antiplatelet antibodies may appear, such as those of chronic idiopathic thrombocytopenia. In this situation, transfused platelets are quickly destroyed, making the transfusion of little value unless the patient is actively bleeding (Chapter 10).

  • Allergic Reactions

    Allergic reactions are the second most common transfusion reaction. They are presumably due to substances in the donor blood to which the recipient is allergic. Symptoms are localized or generalized hives, although occasionally severe asthma or even laryngeal edema may occur. There is usually excellent response to antihistamines or epinephrine.

  • Nonhemolytic Febrile Reactions

    Sometimes called simply (and incorrectly) “febrile reactions,” nonhemolytic febrile reactions occur in about 1% (range, 0.5-5.0%) of all transfusions, more commonly in multiply transfused patients.

    Leukoagglutinin reaction. The most common variety of nonhemolytic febrile reaction and the most common of any transfusion reaction is a febrile episode during or just after transfusion due to patient antibodies against donor leukocytes (leukoagglutinins). Before leukoagglutinins were recognized, these episodes were included among the pyrogenic reactions (discussed later). Fever may be the only symptom but there may be others, such as chills, headache, malaise, confusion, and tachycardia. In a small number of cases a syndrome known as transfusion-related acute lung injury is produced. It includes febrile reaction symptoms plus dyspnea and tachycardia; the chest x-ray film has a characteristic pattern described as numerous hilar and lower lobe nodular infiltrates without cardiac enlargement or pulmonary vessel congestion. In some cases the clinical picture includes marked hypoxemia and also hypotension and thus strongly resembles the adult respiratory distress syndrome. However, symptoms usually substantially improve or disappear in 24-48 hours (81% of patients in one report), but the radiologic abnormalities may persist for several days. Some patients have eosinophilia, but others do not. The syndrome has been called “noncardiac pulmonary edema,” a somewhat unfortunate term since pulmonary edema is not present, although many clinical findings simulate pulmonary edema. Most cases with leukoagglutinin lung involvement follow transfusion of whole blood. At least some of these reactions have been traced to histocompatibility leukocyte antigen (HLA) system incompatibility.

    Most patients produce leukoagglutinins due to previous transfusion or from fetal antigen sensitization in pregnancy. The more transfusions, the greater the likelihood of sensitization.

    Reactions due to leukoagglutinins are usually not life threatening but are unpleasant to the patient, physician, and laboratory. Since clinical symptoms are similar to those of a hemolytic reaction, each febrile reaction must be investigated to rule out RBC incompatibility, even if the patient is known to have leukoagglutinins. Leukoagglutinin reactions may be reduced or eliminated by the use of leukocyte-poor packed RBC, washed RBC, frozen RBC, or best, use of a special leukocyte-removal filter (Chapter 10). There are specialized tests available that are able to demonstrate leukoagglutinin activity, but most laboratories are not equipped to perform them. The diagnosis, therefore, is usually a presumptive one based on history and ruling out hemolytic reaction.

    Washed or frozen RBCs contain fewer white blood cells (WBCs) than the average leukocyte-poor packed cell preparation. However, since washed or frozen RBCs must either be transfused within 24 hours after preparation or be discarded, one must be certain that a need for transfusion exists sufficient to guarantee that the blood will actually be administered before these blood products are ordered. In contrast, leukocyte filtration can be done at the patient’s bedside as part of the transfusion.

    Pyrogenic reactions. These result from contamination by bacteria, dirt, or foreign proteins. One frequently cited study estimates that nearly 2% of donor blood units have some degree of bacterial contamination, regardless of the care taken when the blood was drawn. Symptoms begin during or shortly after transfusion and consist of chills and fever; the more severe cases often have abdominal cramps, nausea, and diarrhea. Very heavy bacterial contamination may lead to shock. Therefore, in a patient with transfusion-associated reaction that includes hypotension, a Gram-stained smear should be made from blood remaining in the donor bag without waiting for culture results.

  • Jaundice in the Newborn or Neonate

    Current consensus criteria indicating pathologic rather than physiologic levels of total bilirubin are the following:

    1. Total bilirubin level over 5 mg/100 ml (88 µmol/L) in the first 24 hours.
    2. Total bilirubin level over 10 mg/100 ml (171 µmol/L) during the second day of life, or an increase of 5 mg/100 ml per day or more thereafter.
    3. Total serum bilirubin level more than 15 mg/100 ml in full-term neonates.
    4. Total serum bilirubin level more than 12 mg/100 ml in premature neonates.
    5. Persistence of jaundice after the first 7 days of life.
    6. Conjugated bilirubin level over 1.5 mg/100 ml at any time.

    A number of conditions in addition to hemolytic disease of the newborn may be associated with elevated bilirubin; some of these conditions include bacterial sepsis, cytomegalovirus or toxoplasma infection, glucose 6-phosphate dehydrogenase deficiency, and resorption of heme from cephalhematoma or extensive bruising. These are uncommon causes of jaundice. Other factors that may increase total bilirubin are the following:

    1. Breast feeding. Although this subject is controversial, a number of studies have found an average increase of about 1.5 mg/100 ml and individual increases up to 2 mg/100 ml or even 3 mg in breast-fed neonates compared to bottle-fed neonates.
    2. Race. In one study, Asian neonates had a 31% incidence of nonphysiologic total bilirubin levels after the first day, compared to 20% in Hispanics, 14% in Europeans, and 9% in African Americans.
    3. Maternal smoking. This is associated with mildly lower infant bilirubin values.
    4. Weight loss and caloric deprivation. These increase serum bilirubin values in both children and adults.
    5. Prematurity. Neonates weighing less than 2,000 gm at birth are on the average about 3 times as likely to develop serum total bilirubin levels over 8 mg/1000 ml and those between 2000-2500 gm are twice as likely to develop such levels as infants with normal birth weight of 2500 gm or more. For bilirubin levels of 14 mg/100 ml, the likelihood is 8 times for the lowest birth rate category and 3 times for the intermediate category.

    Total bilirubin in neonates is almost all nonconjugated, with the conjugated fraction being less than 0.5 mg/100 ml. Elevated conjugated bilirubin level suggests sepsis, liver or biliary tract disease, or congenital abnormality of bilirubin metabolism such as Rotor’s syndrome. However, the level of conjugated bilirubin partially depends on the combination of assay methodology, reagents, and equipment used. Proficiency test surveys such as those of the College of American Pathologists shows that most laboratories have similar results for total bilirubin. For conjugated bilirubin, however, results on a specimen with elevated nonconjugated bilirubin (such as a normal newborn) show normal conjugated bilirubin in some laboratories, significantly elevated conjugated bilirubin in another group of laboratories, and levels between those of the first two groups in a third group of laboratories. Most of the higher and intermediate values were performed on automated chemistry equipment. In my own laboratory, using a well-known automated chemistry analyzer called the Abbott Spectrum, we were in the group with false high values compared to the normal group and the group with intermediate elevated values. Our upper limit of the reference range for conjugated bilirubin on clinically normal adults is 0.4 mg/100 ml. In a study done on newborns with total bilirubin levels ranging from 2-25 mg/100 ml, the Spectrum reported conjugated bilirubin elevations up to 1.2 mg/100 ml when total (nonconjugated) bilirubin was elevated. Surprisingly, once the level of 1.2 mg/100 ml was reached, it would not increase further regardless of further increase in total bilirubin. Therefore, the Spectrum includes some nonconjugated bilirubin in its conjugated bilirubin assay up to a certain point. The majority of laboratories show this phenomenon to some degree. Therefore, for these labs, the reference range upper limit for conjugated bilirubin must be readjusted when the total bilirubin level is elevated.

  • Hemolytic Reactions

    The presence of unexpected alloantibodies (antibodies against red cell antigens) in patient serum found in pretransfusion screening of recipients is 0.7%-1.6%. This averages 9% (range, 6%-36%) in multitransfused patients. Infants less than 4 months old usually do not form alloantibodies against transfused red cell antigens that they lack. After that, age per se does not appear to affect red blood cell (RBC) antigen sensitization. Immunosuppressive therapy can diminish this response. In the case of Rh system D (Rho) antigen, chance of sensitization has some correlation with antigen dose, but this is not exact or linear. Sensitization of D-negative recipients of D-positive cells has ranged from 8% to 70%. Although antibodies to bacterial or many other antigens usually appear in 7-21 days, alloantibodies usually take 3-4 months after transfusion with a minimum (in one study) of 1 month. Once formed, the antibodies remain detectable for variable periods of time, depending to some extent (but not entirely) on the particular antibody. Anti-D is particularly likely to be detectible for many years; anti-C and anti-Kidd are more likely to become nondetectable (50% loss in 5 years in one study). However, nondetectable antibodies can be reactivated by anamnesthic antigen exposure.

    Hemolytic reactions may be caused by either complete or incomplete antibodies. In reactions caused by complete antibodies, such as occur in the ABO blood group system, there is usually intravascular hemolysis. The amount of hemolysis depends on several factors, such as quantity of incompatible blood, antibody titer, and the nature of the antibody involved. However, there is an element of individual susceptibility; some patients die after transfusion of less than 100 ml of incompatible blood, whereas others survive transfusion of several times this amount. The direct Coombs’ test result is often positive, but this depends on whether all the RBCs attacked by the complete antibody have been lysed, whether more antibody is produced, and, to some extent, on how soon the test is obtained. If the sample is drawn more than 1 hour after the ABO transfusion reaction is completed, the chance of a positive direct Coombs’ test result is much less. A Coombs’ reagent acting against both gamma globulin and complement (broad spectrum) is needed; most laboratories now use this type routinely. Free hemoglobin is released into the plasma from lysed RBCs and is carried to the kidneys, where it is excreted in the urine. Some of the intravascular hemoglobin is converted to bilirubin in the reticuloendothelial system so that the serum nonconjugated (indirect) bilirubin level usually begins to increase. In reactions caused by incomplete antibodies, such as the Rh system, there is sequestration of antibody-coated cells in organs such as the spleen, with subsequent breakdown by the reticuloendothelial system. With incomplete antibody reactions, RBC breakdown is extravascular rather than intravascular; in small degrees of reaction, plasma free hemoglobin levels may not rise, although indirect bilirubin levels may eventually increase. In more extensive reactions, the plasma hemoglobin level is often elevated, although the elevation is sometimes delayed in onset. The direct Coombs’ test should be positive in hemolytic reactions due to incomplete antibodies (unless all affected RBCs have been destroyed).

    Undesirable effects of blood or blood product transfusion

    RECIPIENT REACTION TO DONOR ANTIGENS ON DONOR CELLS
    Clinical types of reactions
    Hemolytic reaction
    Nonhemolytic febrile reactions
    Allergic reactions
    Cytopenic reactions
    Tissue-organimmunologic reactions
    Anaphylactic reactions

    ANTIGEN GROUPS INVOLVED
    ABO system
    Rh and minor blood groups
    Histocompatibility leukocyte antigen (HLA) system
    Platelet antigens

    INFECTIONS
    Hepatitis viruses
    Human immunodeficiency virus-1 and 2 (HIV-1 and 2)
    Human T-cell lymphotropic virus-1 and 2 (HTLV-1 and II)
    Cytomegalovirus (CMV)
    Epstein-Barr virus
    Syphilis
    Malaria
    Other

    OTHER TRANSFUSION PROBLEMS
    Citrate overload
    Hyperkalemia
    Depletion of coagulation factors
    Depletion of platelets
    Transfused blood temperature
    Donor medications

    Hemolytic transfusion reactions are usually caused by incompatible blood but are occasionally caused by partial hemolysis of the RBCs before administration, either before leaving the blood bank or just before transfusion if the blood is warmed improperly. Analysis of fatal cases of hemolytic transfusion reaction reported to the Food and Drug Administration in 1976-1983 shows that almost 65% were due to problems of mistaken identity and about 9% were due to clerical error. Only about 18% were due to error while performing blood bank tests. Frequent mistakes included transfusion into one patient of blood meant for another patient, obtaining a recipient crossmatch specimen from the wrong patient, mixup of patient crossmatch specimens in the blood bank, and transcription of data belonging to one patient onto the report of another.

    The great majority of hemolytic transfusion reactions occur during transfusion. Symptoms and laboratory evidence of hemolysis usually are present by the time transfusion of the incompatible unit is completed, although clinical symptoms or laboratory abnormalities may sometimes be delayed for a few minutes or even a few hours. Occasionally, hemolytic reactions take place after completion of transfusion (delayed reaction). In one kind of delayed reaction, the reaction occurs 4-5 days (range, 1-7 days) following transfusion and is due to anamnestic stimulation of antibodies that were already present but in very low titer. The Kidd (Jk) system is frequently associated with this group. The intensity of the reaction may be mild or may be clinically evident, but most are not severe. In a second kind of delayed reaction, the reaction occurs several weeks after transfusion and is due to new immunization by the transfused RBC antigens with new antibody production. This type of reaction is usually mild and subclinical. In both types of reaction, and occasionally even in an immediate acute reaction, a hemolytic reaction may not be suspected and the problem is discovered accidentally by a drop in hemoglobin level or by crossmatching for another transfusion.

    Symptoms of hemolytic transfusion reaction include chills, fever, and pain in the low back or legs. Jaundice may appear later. Severe reactions lead to shock. Renal failure (acute tubular necrosis) is common due either to shock or to precipitation of free hemoglobin in the renal tubules. Therefore, oliguria develops, frequently accompanied by hemoglobinuria, which is typically manifest by red urine or nearly black “coffee ground” acid hematin urine color.

    Tests for hemolytic transfusion reaction

    These tests include immediate rechecking and comparison of patient identification and the unit or units of transfused donor blood. Tubes of anticoagulated and nonanticoagulated blood should be drawn. The anticoagulated tube should be centrifuged immediately and the plasma examined visually for the pink or red color produced by hemolysis. A direct Coombs’ test and crossmatch recheck studies should be performed as soon as possible. A culture and Gram stain should be done on the remaining blood in the donor bag.

    Plasma free hemoglobin. The presence of free hemoglobin in plasma is one of the most valuable tests for diagnosis of acute hemolytic transfusion reaction. Most cases of severe intravascular hemolysis due to bivalent antibodies such as the ABO group produce enough hemolysis to be grossly visible. (Artifactual hemolysis from improperly drawn specimens must be ruled out.) If chemical tests are to be done, plasma hemoglobin is preferred to serum hemoglobin because less artifactual hemo-lysis takes place before the specimen reaches the laboratory. Although hemolytic reactions due to incomplete antibodies (such as Rh) have clinical symptoms similar to those of ABO, direct signs of hemolysis (e.g., free plasma hemoglobin) are more variable or may be delayed for a few hours, although they become abnormal if the reaction is severe. Another drawback in the interpretation of plasma hemoglobin values is the effect of the transfusion itself. The older erythrocytes stored in banked blood die during storage, adding free hemoglobin to the plasma. Therefore, although reference values are usually considered to be in the range of 1-5 mg/100 ml, values between 5 and 50 mg/100 ml are equivocal if stored blood is given. Hemolysis is barely visible when plasma hemoglobin reaches the 25-50 mg/100 ml range. Frozen blood has a greater content of free hemoglobin than stored whole blood or packed cells and may reach 300 mg/100 ml.

    Additional tests. If visual inspection of patient plasma does not suggest hemolysis and the direct Coombs’ test result is negative, additional laboratory tests may be desirable to detect or confirm a hemolytic transfusion reaction. Procedures that may be useful include serum haptoglobin measurement, both immediately and 6 hours later; serum nonconjugated (“indirect”) bilirubin measurement at 6 and 12 hours; and urinalysis.

    A pretransfusion blood specimen, if available, should be included in a transfusion reaction investigation to provide baseline data when the various tests for hemolysis are performed.

    Urinalysis. Urine can be examined for hemoglobin casts, RBCs, and protein and tested for free hemoglobin. RBCs and RBC casts may be found in hemolytic reactions since the kidney is frequently injured. If no hematuria or free hemoglobin is found, this is some evidence against a major acute hemolytic transfusion reaction. Abnormal findings are more difficult to interpret unless a pretransfusion urinalysis result is available for comparison, since the abnormality might have been present before transfusion. In addition, abnormal urine findings could result from hypoxic renal damage due to an episode of hypotension during surgery rather than a transfusion reaction.

    Hemoglobin passes through the glomerular filter into the urine when the plasma hemoglobin level is above 125 mg/100 ml. Therefore, since hemolysis should already be visible, the major need for urine examination is to verify that intravascular hemolysis occurred rather than artifactual hemolysis from venipuncture or faulty specimen processing. Conversely, unless the plasma free hemoglobin level is elevated, urine hemoglobin may represent lysed RBCs from hematuria unrelated to a hemolytic reaction. However, if sufficient time passes, the serum may be cleared of free hemoglobin while hemoglobin casts are still present in urine. Hemosiderin may be deposited in renal tubule cells and continue to appear in the urine for several days.

    Serum haptoglobin. Haptoglobin is an alpha globulin that binds free hemoglobin. Two types of measurements are available: haptoglobin-binding capacity and total haptoglobin. Binding capacity can be measured by electrophoresis or chemical methods. Total haptoglobin is usually estimated by immunologic (antibody) techniques; a 2-minute slide test is now commercially available. The haptoglobin-binding capacity decreases almost immediately when a sufficient quantity of free hemoglobin is liberated and remains low for 2-4 days. Stored banked blood, fortunately, does not contain enough free hemoglobin to produce significant changes in haptoglobin-binding capacity. The total haptoglobin level decreases more slowly than the binding capacity after onset of a hemolytic reaction and might not reach its lowest value until 6-8 hours later. Haptoglobin assay is much less helpful when frozen RBCs are transfused because of the normally increased free hemoglobin in most frozen RBC preparations. Bilirubin determinations are not needed if haptoglobin assay is done the same day.

    In common with other laboratory tests, serum haptoglobin levels may be influenced by various factors other than the one for which the test is ordered. Haptoglobin levels may be decreased by absorption of hemoglobin from an extravascular hematoma and also may be decreased in severe liver disease, hemolytic or megaloblastic anemia, estrogen therapy, and pump-assisted open-heart cardiac surgery. Haptoglobin is one of the body’s “acute-phase reactant” proteins that are increased by infection, tissue injury or destruction, widespread malignancy, and adrenocorticosteroid therapy. Therefore, a mild decrease in haptoglobin level could be masked by one of these conditions. In order to aid interpretation, it is helpful to perform the haptoglobin assay on a pretransfusion serum specimen as well as the posttransfusion specimen.

    Sensitivity of tests in hemolytic reaction. Data regarding frequency of test abnormality in transfusion reaction are difficult to obtain. In one series, comprising predominantly hemolytic reactions not due to ABO antibodies, free hemoglobin was detected in plasma or urine in 88% of cases involving immediate reactions and in 52% of cases involving incomplete antibodies or delayed reactions. Serum haptoglobin levels were decreased in 92% of the patients in whom it was assayed. Various factors influence this type of data, such as the amount of incompatible blood, the antibody involved, the time relationship of specimen to onset of reaction, and the test method used.

    Nursing station action in possible hemolytic reaction

    Transfusion should be stopped at the first sign of possible reaction and complete studies done to recheck compatibility of the donor and recipient blood. If these are still satisfactory, and if results of the direct Coombs’ test and the studies for intravascular hemolysis are negative, a different unit can be started on the assumption that the symptoms were pyrogenic rather than hemolytic. Whatever unused blood remains in the donor bottle, the donor bottle pilot tube, and a new specimen drawn from the patient must all be sent to the blood bank for recheck studies. Especially dangerous situations exist in transfusion during surgery, where anesthesia may mask the early signs and symptoms of a reaction. Development during surgery of a marked bleeding or oozing tendency at the operative site is an important danger signal. A hemolytic transfusion reaction requires immediate mannitol or equivalent therapy to protect the kidneys.

    Hemolytic disease of the newborn (HDN)

    The other major area where blood banks meet blood group hemolytic problems is in hemolytic disease of the newborn (HDN). It may be due to ABO, Rh, or (rarely) minor group incompatibility between fetal and maternal RBCs. ABO and the Rh antigen D (Rho) are by far the most common causes. The Rh antigen c and the blood group Kell antigen are next most important. Hemolytic disease of the newborn results from fetal RBC antigens that the maternal RBCs lack. These fetal RBC antigens provoke maternal antibody formation of the IgG type when fetal RBCs are introduced into the maternal circulation after escaping from the placenta. The maternal antibodies eventually cross the placenta to the fetal circulation and attack the fetal RBCs.

    Hemolytic disease of the newborn due to Rh. Hemolytic disease of the newborn due to Rh incompatibility varies in severity from subclinical status, through mild jaundice with anemia, to the dangerous and often fatal condition of erythroblastosis fetalis. The main clinical findings are anemia and rapidly developing jaundice. Reticulocytosis over 6% accompanies the anemia, and the jaundice is mainly due to unconjugated (indirect) bilirubin released from the reticuloendothelial sequestration and destruction of RBCs. The direct Coombs’ test result is positive. In severe cases there are usually many nucleated RBCs in the peripheral blood. Jaundice is typically not present at birth except in very severe cases (since the mother excretes bilirubin produced by the fetus) but develops several hours later or even after 24 hours in mild cases. Diseases that cause jaundice in the newborn, often accompanied by anemia and sometimes a few peripheral blood nucleated RBCs, include septicemia, cytomegalic inclusion disease, toxoplasmosis, and syphilis. Physiologic jaundice of the newborn is a frequent benign condition that may be confused with hemolytic disease or vice versa. There is, however, no significant anemia. A normal newborn has a (average) hemoglobin value of 18 gm/100 ml, and a value less than 15 gm/100 ml (150 g/L) indicates anemia. Anemia may be masked if heelstick (capillary) blood is used, since neonatal capillary hemoglobin values are higher than venous blood values.

    Hemolytic disease due to Rh incompatibility occurs in an Rh-negative mother whose fetus is Rh positive. Usually, the mother and fetus are ABO compatible. The mother develops antibodies against the RBC Rh antigen after being exposed to Rh-positive RBCs. This may occur due to pregnancy, abortion, ectopic pregnancy, amniocentesis (or other placental trauma), blood transfusion with Rh-positive RBCs, or transfusion of certain RBC-contaminated blood products such as platelets. The most common maternal contact with Rh-positive RBCs occurs during pregnancy when fetal RBCs escape through the placenta into the maternal circulation. This may happen at any time after the 16th week of pregnancy, and both the quantity of cells and the frequency of exposure increase until delivery. The largest single dose of fetal RBCs occurs during delivery. Not all mothers have detectable fetal RBCs in their circulation, and of those who do, not all become sensitized during any one pregnancy. There is approximately a 10%-13% risk of sensitization in previously nonsensitized Rh-incompatible pregnancies. When fetal-maternal ABO incompatibility (with the mother being group O) is present, the usual risk for Rh sensitization is decreased, presumably because sufficient fetal cells are destroyed that the stimulus for sensitization is reduced below the necessary level.

    The first child is usually not affected by Rh hemolytic disease if the mother has not been exposed to Rh-positive RBCs before pregnancy, and full sensitization usually does not develop until after delivery in those mothers who become sensitized. However, occasional firstborn infants are affected (5%-10% of HDN infants) either because of previous maternal exposure (e.g., a previous aborted pregnancy) or because of unusually great maternal susceptibility to Rh stimulus during normal pregnancy. Once maternal sensitization takes place, future exposure to Rh antigen, as during another pregnancy with an Rh-positive fetus, results in maternal antibody production against the Rh antigen, which can affect the fetus.

    Current recommendations of the American College of Obstetricians and Gynecologists (ACOG) are that every pregnant woman should have ABO and Rh typing and a serum antibody screen as early as possible during each pregnancy. If results of the antibody screen are negative and the mother is Rh positive, the antibody screen would not have to be repeated before delivery. Theoretically, if the mother is Rh positive, or if the mother is Rh negative and the father is also Rh negative, there should be no risk of Rh-induced fetal disease. However, the antibody screen is still necessary to detect appearance of non-Rh antibodies. If results of the antibody screen are negative and the mother is Rh negative, the father should be typed for Rh and the antibody screen should be repeated at 28 weeks’ gestation. If the antibody screen results are still negative, a prophylactic dose of Rh immune globulin is recommended (discussed later). If the antibody screen detects an antibody, subsequent testing or action depends on whether the antibody is Rh or some other blood group and whether or not this is the first pregnancy that the antibody was detected. The father should be tested to see if he has the antigen corresponding to the antibody. If the antibody is one of the Rh group, antibody titers should be performed every 2 weeks. Titers less than 1:16 suggest less risk of fetal hazard. Titer equal to or greater than 1:16 is usually followed by amniocentesis (as early as 24 weeks’ gestation) for spectrophotometric examination of amniotic fluid bilirubin pigment density. The greater the pigment density the greater the degree of fetal RBC destruction. Antibodies that are not Rh are managed by amniotic fluid examination without antibody titers, since there is inadequate correlation of titers to fetal outcome.

    Rh immune globulin. Studies have now proved that nearly all nonsensitized mothers with potential or actual Rh incompatibility problems can be protected against sensitization from the fetus by means of a “vaccine” composed of gamma globulin with a high titer of anti-Rh antibody (RhIg). This exogenous antibody seems to prevent sensitization by destroying fetal RBCs within the mother’s circulation and also by suppressing maternal antibody production. Abortion (spontaneous or induced), ectopic pregnancy, amniocentesis, and platelet or granulocyte transfusions (which may be contaminated with Rh-positive RBCs) may also induce sensitization and should be treated with RhIg. Current ACOG recommendations are to administer one unit (300 µg) of RhIg to all Rh-negative women at 28 weeks’ gestation and a second unit within 72 hours after delivery. A minidose of 50 µg is often used rather than the standard dose when abortion occurs before 12 weeks’ gestation. Although 72 hours after delivery is the recommended time limit for RhIg administration, there is some evidence that some effect may be obtained up to 7 days after delivery. Untreated Rh-negative women have about a 10% (range, 7%-13%) incidence of Rh antibody production (sensitization). When RhIg is administered postpartum it reduces incidence to about 1%-2%, and antepartum use combined with postpartum use reduces the incidence to about 0.1%-0.2%. RhIg will produce a positive Rh antibody screening test result if present in sufficient titer, so that antibody screening should be performed before administration rather than after. The half-life of exogenous gamma globulin (which applies to RhIg) is about 25 days (range 23-28 days). Injected RhIg may be detected in maternal blood after intramuscular injection within 24-48 hours with a peak at about 5 days, and is often still detectable for 3 months (sometimes as long as 6 months). Detection after 6 months postpartum suggests patient sensitization (failure of the RhIg). There have been a few reports that some Rh-positive infants whose mother received antepartum RhIg had a weakly positive direct Coombs’ test result at birth due to the RhIg. The blood bank should always be informed if RhIg has been given antepartum to properly interpret laboratory test results. The standard dose of 300 µg of RhIg neutralizes approximately 15 ml of Rh-positive RBCs (equivalent to 30 ml of fetal whole blood). In some patients the RBCs received from the fetus may total more than 15 ml.

    There are several methods used to quantitate fetal RBCs in the maternal circulation, none of which are considered ideal. The Kleihauer acid elution test is a peripheral smear technique that stains the hemoglobin F (Hb F) of fetal RBCs. Other tests that have been used are the weak D (Du) test read microscopically (to detect the mixed field agglutination of the relatively small number of fetal RBCs involved) and the RhIg crossmatch technique. None of these three procedures has proved adequately sensitive. For example, a proficiency survey in 1980 found that about 10% of the laboratories using acid elution or microscopic Du techniques failed to detect the equivalent of 30 ml of fetal RBCs (twice the significant level). When a normal blood sample was tested, about 10% of those using microscopic Du obtained false positive results, and those using acid elution had 40% or more false positive results (especially with one commercial adaptation of the acid elution technique called Fetaldex). Also, false positive acid elution test results can be produced if increased maternal Hb F is present, as seen in beta thalassemia minor and in hereditary persistence of fetal hemoglobin. Newer procedures, such as the erythrocyte rosette test, have proved much more sensitive and somewhat more reproducible. However, the rosette test is not quantitative, so that screening is done with the rosette test (or equivalent) and a positive result is followed by quantitation with the acid elution procedure. If the quantitative test for fetal RBCs indicates that more than 15 ml is present, additional RhIg should be administered. If this is not done, failure rates for RhIg of 10%-15% have been reported; if it is done, the failure rate is approximately 2%. The current American Association of Blood Banks (AABB) recommendation is to administer twice the dose of RhIg indicated by formulas, depending on the percent of fetal RBCs detected by acid elution methods. This is done because of variation above and below the correct result found on proficiency surveys.

    ABO-induced hemolytic disease of the newborn

    Fifty percent or more of HDN is due to ABO incompatibility between mother and fetus. There usually is no history of previous transfusion. Most cases are found in mothers of blood group O with group A or B infants. Infant anti-A and anti-B production begins between 3 and 6 months after birth. Until then, ABO antibodies in the infant’s serum originate from the mother. If the mother possesses antibodies to the A or B RBC antigens of the fetus, hemolytic disease of the fetus or newborn may result, just as if the fetus or newborn had received a transfusion of serum with antibodies against his or her RBCs. Nevertheless, although 20%-25% of all pregnancies display maternal-fetal ABO incompatibility, only about 40% of these infants develop any evidence of hemolytic disease. In those who do, the condition is usually clinically milder than its counterpart caused by Rh incompatibility with only 4%-11% displaying clinical evidence of disease. There usually are no symptoms at birth. Some infants, however, will suffer cerebral damage or even die if treatment is not given. Therefore, the diagnosis of ABO disease and its differential diagnosis from the other causes of jaundice and anemia in the newborn are of great practical importance.

    There are two types of ABO antibodies: the naturally occurring complete saline-reacting type discussed earlier and an immune univalent (incomplete) type produced in some unknown way to fetal A or B antigen stimulation. In most cases of clinical ABO disease the mother is group O and the infant is group A or B. The immune anti-A or anti-B antibody, if produced by the mother, may cause ABO disease because it can pass the placenta. Maternal titers of 1:16 or more are considered dangerous. The saline antibodies do not cross the placenta and are not significant in HDN.

    Results of the direct Coombs’ test done on the infant with ABO hemolytic disease are probably more often negative than positive, and even when positive the test tends to be only weakly reactive. Spherocytes are often present but the number is variable. Good evidence for ABO disease is detection of immune anti-A or anti-B antibodies in the cord blood of a newborn whose RBCs belong to the same blood group as the antibody. Detection of these antibodies only in the serum of the mother does not conclusively prove that ABO disease exists in the newborn.

    Laboratory tests in hemolytic disease of newborns (HDN)

    When an infant is affected by Rh-induced HDN, the result of the direct Coombs’ test on cord blood is nearly always positive. In ABO-induced HDN the direct Coombs’ test result on cord blood is frequently (but not always) positive. The direct Coombs’ test result on infant peripheral blood is usually positive in Rh-induced disease but is frequently negative in ABO-induced disease, especially when done more than 24 hours after delivery. The direct Coombs’ test should be performed in all cases of possible HDN, because incomplete antibodies occasionally coat the surface of fetal RBCs to such an extent as to interfere with proper Rh typing. Cord blood bilirubin is usually increased and cord blood hemoglobin is decreased in severe HDN. There is disagreement whether cord blood hemoglobin level has a better correlation with disease severity than the cord blood bilirubin or infant venous or capillary hemoglobin level. Infant hemoglobin levels tend to be higher than cord hemoglobin levels if blood from the cord and placenta is allowed to reach the infant after delivery.

    Laboratory criteria for hemolytic disease of the newborn

    Infants with HDN can frequently be saved by exchange transfusion. Commonly accepted indications for this procedure are the following:

    1. Infant serum indirect bilirubin level more than 20 mg/100 ml (342 µmol/L) or, in considerably premature or severely ill infants, 15 mg/100 ml (257 µmol/L).
    2. Cord blood indirect bilirubin level more than 3 mg/100 ml (51 µmol/L) (some require 4 mg/100 ml).
    3. Cord blood hemoglobin level less than 13 gm/dl (130 g/L)(literature range, 8-14 gm/100 ml).
    4. Maternal Rh antibody titer of 1:64 or greater, although this is not an absolute indication if the bilirubin does not rise very high.

    Bilirubin levels in hemolytic disease of the newborn. Most infants with HDN can be treated effectively enough that exchange transfusion is not needed. The level of infant bilirubin is generally used as the major guideline for decision and to monitor results of other treatment such as phototherapy. An infant total bilirubin level of 12-15 mg/100 ml (205 µmol/L) depending to some extent on the clinical situation (degree of prematurity, presence of anemia or infection, severity of symptoms, etc.) is the most commonly accepted area at which therapy is begun. However, there is surprising variation in the levels quoted in various medical centers. Rarely, kernicterus has been reported in seriously ill infants at bilirubin levels near 10 mg/100 ml (and in one case even as low as 6 mg/100 ml). The rapidity with which the bilirubin level rises is also a factor, as noted previously. To further complicate matters, autopsy studies have shown yellow staining of brain tissue typical of kernicterus in some infants who did not have clinical symptoms of kernicterus.

    Bilirubin levels and kernicterus. The most feared complication of Rh-induced hemolytic disease of the newborn is kernicterus, defined as bilirubin staining of the central nervous system (CNS) basal ganglia with death or permanent neuro-logic or mental abnormalities. When this syndrome was first studied in the 1950s, Rh-induced hemolytic disease was the usual etiology, and the nonconjugated bilirubin level of 20 mg/100 ml in term infants (15 mg/dl in premature infants) was established as the level at which the kernicterus syndrome was most likely to develop and thus the level at which exchange transfusion was required. It was also reported that various other factors, such as acidosis, respiratory distress, infection, and very low birth weight could be associated with the kernicterus syndrome at bilirubin levels less than 15 mg/100 ml (several case reports included a few infants whose total bilirubin level was as low as 9-10 mg/100 ml). Eventually the nonconjugated bilirubin level rather than infant symptoms became the center of attention (however, since neonatal bilirubin except in rare cases is almost all nonconjugated, total bilirubin level is routinely assayed instead of nonconjugated bilirubin). As time went on, the advent of RhIg therapy markedly reduced Rh hemolytic disease, and neonatal jaundice became over 90% nonhemolytic. More recent studies have questioned the relationship between total bilirubin level and the kernicterus syndrome. Although phototherapy can reduce total bilirubin levels, there is some question whether in fact this can prevent kernicterus. Therefore, in the early 1990s there is very low incidence of the kernicterus syndrome, the mechanisms and pathologic basis for this syndrome is uncertain, the relationship and interpretation of nonconjugated or total bilirubin levels is being questioned, and the classic guidelines for therapy are being disputed. Nevertheless, while the situation is unclear, the majority of investigators appear to be using 15 mg/100 ml as the level to begin phototherapy (less if the infant is premature and severely ill) and 20 mg/100 ml as the level to consider intensive therapy (possibly, but not necessarily including exchange transfusion). Exchange transfusion appears to be reserved more for infants with severe hemolytic disease; for example, Rh or Kell incompatibility, neonatal glucose 6-phosphate dehydrogenase hemolysis, and sepsis.

    In addition, there are certain technical problems involving bilirubin assay. Phototherapy breaks down nonconjugated bilirubin into nontoxic bilirubin isomers, which, however, are measured and included with unconjugated bilirubin in standard bilirubin assays. Finally, it must be mentioned that bilirubin is one of the least accurate of frequently or routinely performed chemistry assays, with proficiency surveys consistently showing between-laboratory coefficients of variation of 10% or more. To this is added variances due to specimens obtained under different conditions (venous vs. capillary or heelstick), state of infant hydration, and differences between laboratories because of different methodologies. At present, the bilirubin measurement within the capability of most laboratories that is considered to best correlate with clinical kernicterus is unconjugated (indirect) bilirubin. In HDN, most of the bilirubin will be unconjugated. Therefore, it usually is sufficient to obtain total bilirubin levels in order to follow the patient’s progress. If there is a question about the diagnosis, one request for conjugated/unconjugated fractionation is sufficient. A definite consideration is the additional blood needed to assay both the conjugated and unconjugated fractions since large specimens may be difficult to obtain from an infant heel puncture.

    Albumin-bound bilirubin. Unconjugated bilirubin is presumed to be the cause of kernicterus. However, unconjugated or total bilirubin levels do not always correlate well with development of kernicterus. In the 1980s there was interest in measurement of free nonconjugated bilirubin and bilirubin-binding capacity. Most of the unconjugated bilirubin in serum is tightly bound to serum albumin, with a small portion being loosely bound (“free”). Since the free portion rather than the tightly bound portion theoretically should be the most important element in kernicterus, various methods have been devised to assay free unconjugated bilirubin rather than total bilirubin. Until approximately 1980, direct measurement was difficult, and most attention was given to indirect methods, chiefly estimation of the bilirubin-binding capacity of albumin. In general, when more bilirubin binds to albumin, the residual binding capacity becomes smaller and less binding of serum free bilirubin takes place. Therefore, free bilirubin levels are more likely to increase. Several methods have been proposed to measure albumin-binding capacity; the most popular involved a Sephadex resin column. Sephadex resin competes with albumin for loosely bound bilirubin. When unconjugated bilirubin levels exceed the bilirubin-binding capacity of albumin, the excess binds to the sephadex column. There are conflicting reports in the literature on the value of the albumin-binding capacity assay. Some reports indicated that it was very helpful; others found that results from individual patients were either too often borderline or did not correlate sufficiently well with the clinical picture. More recently, an instrument called the hematofluorometer that measures total bilirubin-binding capacity (TBBC) has become available. Total bilirubin-binding capacity as measured by the hematofluorometer is reported to correlate well with unbound bilirubin. Nevertheless, there does not appear to be convincing evidence that albumin-binding capacity or free bilirubin measurements have shown clear-cut superiority over traditional guidelines. In summary, although a total bilirubin level of 20 mg/100 ml is a reasonably good cutoff point for substantial risk of kernicterus in full-term infants, no adequate cutoff point has been found for sick premature infants; and consideration of risk factors such as sepsis, acidosis, pulmonary distress, hypothermia, hypoalbuminemia, or bilirubin-binding capacity has not produced totally reliable criteria for determining whether to use therapy or not.

    Role of amniocentesis in hemolytic disease of the newborn. Amniocentesis has been advocated in selected patients as a means of estimating risk of severe HDN while the fetus is still in utero. A long needle is introduced into the amniotic fluid cavity by a suprapubic puncture approach in the mother. The amniotic fluid is subjected to spectro photometric estimation of bilirubin pigments. Markedly increased bilirubin pigment levels strongly indicate significant degrees of hemolytic disease in the fetus. If necessary, delivery can be induced prematurely once the 32nd week of gestation has arrived. Before this, or if the fetus is severely diseased, intrauterine exchange transfusion has been attempted using a transabdominal approach. The indications for amniocentesis are development of significant titer of Rh antibody in the mother or a history of previous erythroblastosis. Significant maternal antibody Rh titers do not always mean serious fetal Rh disease, but absence of significant titer nearly always indicates a benign prognosis. Also, if initial amniocentesis at 32 weeks does not suggest an immediately dangerous situation, even though mild or moderate abnormalities are present, a fetus can be allowed to mature as long as possible (being monitored by repeated studies) to avoid the danger of premature birth.

  • Other Procedures Relevant To Transfusion Therapy

    Apheresis
    Apheresis is a technique in which blood is withdrawn from a donor, one or more substances are removed from the blood, and the blood is then returned to the donor. Most present-day apheresis equipment is based on separation of blood components by differential centrifugation. Apheresis has two major applications. One is the removal of certain blood components (e.g., platelets) to be used for transfusion into another patient. Apheresis has permitted collection of blood components in greater quantity and more frequently than ordinary phlebotomy. In addition, when all components come from the same donor, the risk of hepatitis is less than if blood from multiple donors is used. This technique is the major source of many rare blood group antibodies and certain blood components. The second major use of apheresis is the direct therapeutic removal of harmful substances or blood components from a patient. The most common application is removal of abnormal proteins from serum by plasmapheresis in patients with the hyperviscosity syndrome (associated with Waldenstцm’s macroglobulinemia or myeloma). Apheresis has also been used to remove immune complexes from patients with various disorders associated with autoimmune disease. However, use of apheresis in many of these conditions is considered experimental.

    Blood transfusion filters

    For many years it has been the custom during blood transfusion to place a filter with a 170-µ pore size between the blood donor bag and the patient. This trapped any blood clots large enough to be visible that might have formed in the donor bag. In the 1960s it was recognized that nonvisible clots or microaggregates of platelets, fibrin, degenerating leukocytes, and other debris frequently were present in stored bank blood and could produce microembolization with pulmonary and cerebral symptoms. The most severe condition was pulmonary insufficiency associated with open-heart surgery or massive blood transfusion for severe trauma. It was found that microfilters with pore sizes of 20-40 µ could trap most of these microaggregates. Some publications advocated using such a filter for every transfusion (the filter can accept 5-10 units of whole blood or packed cells before it must be replaced). Others believe that transfusions limited to one or two units do not subject the lungs to sufficient embolized material to necessitate use of a microfilter. Originally there was a flow rate problem with the 20 µ filters, but newer models have better flow characteristics. A substantial number of platelets are trapped by filter sizes less than 40 µ. However, blood more than 2 days old does not contain viable platelets.

    Blood volume measurement

    Blood volume measurement is useful in certain circumstances: (1) to differentiate anemia due to hemodilution from anemia due to RBC deficiency, (2) to differentiate polycythemia from dehydration, and (3) to quantitate blood volume for replacement or for therapeutic phlebotomy purposes. Blood volume measurement is discussed in this chapter since the most frequent indication for transfusion therapy is to replace depleted blood volume. This most commonly arises in association with surgery or from nonsurgical blood loss, acute or chronic. Immediately after an acute bleeding episode, the Hb and hematocrit values are unchanged (because whole blood has been lost), even though the total blood volume may be greatly reduced, even to the point of circulatory collapse (shock). With the passage of time, extracellular fluid begins to diffuse into the vascular system to partially restore total blood volume. Since the hematocrit is simply the percentage of RBCs compared with total blood volume (total blood volume being the RBC mass plus the plasma volume), this dilution of the blood by extracellular fluid means that the hematocrit value eventually decreases, even while total blood volume is being increased (by extracellular fluid increasing plasma volume). Hemodilution (and thus the drop in hematocrit) may be hastened if the patient has been receiving intravenous (IV) fluids. Serial hematocrit determinations (once every 2-4 hours) may thus be used as a rough indication of blood volume changes. It usually takes at least 2 hours after an acute bleeding episode for a significant drop in hematocrit value to be demonstrated. Sometimes it takes longer, even as long as 6-12 hours. The larger the extracellular blood loss, the sooner a significant hematocrit value change (> 2%) is likely to appear.

    Previous dehydration, a low plasma protein level, or both will tend to delay a hematocrit drop. Besides the uncertainty introduced by time lag, other conditions may affect the hematocrit value and thus influence its interpretation as a parameter of blood volume. Anemias due to RBC hemolysis or hematopoietic factor deficiencies such as iron may decrease RBC mass without decreasing plasma or total blood volume. Similarly, plasma volume may be altered in many situations involving fluid and electrolyte imbalance without changing the RBC mass. Obviously, a need exists for accurate methods of measuring blood volume.

    Blood volume methods. The first widely used direct blood volume measurement technique was Evan’s Blue dye (T-1824). After IV injection of the dye, the amount of dilution produced by the patient’s plasma was measured, and from this the plasma volume was calculated. At present, radioisotopes are the procedure of choice. These methods also are based on the dilution principle. Chromium 51 can be used to tag RBCs; a measured amount of the tagged RBCs is then injected into the patient. After equilibration for 15 minutes, a blood sample is obtained and radioactivity is measured. Since the tagged RBCs have mixed with the patient’s RBCs, comparison of the radioactivity in the patient’s RBCs with the original isotope specimen that was injected reveals the amount that the original isotope specimen has been diluted by the patient’s RBCs; thus, the patient’s total RBC mass (RBC volume) may be calculated. If the RBC mass is known, the plasma volume and total blood volume may be derived using the hematocrit value of the patient’s blood. Another widely used method uses serum albumin labeled with radioactive iodine (RISA). This substance circulates in the plasma along with the other plasma proteins. Again, a measured amount is injected, a blood sample is withdrawn after a short period of equilibration, and the dilution of the original injected specimen is determined by counting the radioactivity of the patient’s plasma. Plasma volume is provided by RISA measurement; RBC volume must be calculated using the patient’s hematocrit value. There is no doubt that isotope techniques are much more accurate than the hematocrit value for estimating blood volume. Nevertheless, there are certain limitations to isotope techniques in general and specific limitations to both 51Cr and RISA (see box below).

    Assay problems. The main drawback of blood volume techniques is the lack of satisfactory reference values. Attempts have been made to establish reference values for males and females in terms of height, weight, surface area, or lean body mass. Unfortunately, when one tries to apply these formulas to an individual patient, there is never any guarantee that the patient fits whatever category of normal persons that the formula was calculated from. The only way to be certain is to have a blood volume measurement from a time when the patient was healthy or before the bleeding episode. Unfortunately, this information is usually not available. Another drawback is the fact that no dilution method can detect bleeding that is going on during the test itself. This is so because whole blood lost during the test contains isotope in the same proportion as the blood remaining in the vascular system (a diminished isotope dose in a

    Factors That Can Adversely Affect the Conditions Necessary for Accurate Blood Volume Results
    1. All isotope must be delivered into the fluid volume being measured:
    Extravasation outside vein (falsely increased result)

    2.Need uniform mixing of isotope throughout fluid volume being measured:

    Capillary sequestration in shock
    Slow mixing in congestive heart failure
    Focal sequestration in splenomegaly

    3. Need uniform mixing of isotope throughout fluid volume being measured:
    Male vs. female
    Obesity or malnourishment

    diminished blood volume) in contrast to the situation that would prevail if bleeding were not going on, when the entire isotope dose would remain in a diminished blood volume. Fortunately, such active bleeding would have to be severe for test results to be materially affected. Another problem is dependence on hematocrit value when results for RISA are used to calculate RBC mass or data from 51Cr are used to obtain plasma volume. It is well established that hematocrit values from different body areas or different-sized vessels can vary considerably, and disease may accentuate this variation. The venous hematocrit value may therefore not be representative of the average vascular hematocrit value.

    Single-tag versus double-tag methods. All authorities in the field agree that blood volume determination combining independent measurement of RBC mass by 51Cr and plasma volume by RISA is more accurate than the use of either isotope alone. Nevertheless, because both isotopes must be counted separately with special equipment, most laboratories use only a single isotope technique. Most authorities concede that 51Cr has a slightly better overall accuracy than RISA, although some dispute this strongly. However, most 51Cr techniques call for an extra venipuncture to obtain RBCs from the patient for tagging, plus an extra 30-minute wait while the actual tagging of the cells takes place. In addition, tagged RBCs (and their radioactivity) remain in the circulation during the life span of the cells. The main advantages of RISA are the need for one less venipuncture than 51Cr and the fact that RISA procedures can be done in less than half the time of 51Cr studies. The error with RISA blood volume has reached 300 ml in some studies, although most determinations come much closer to double-isotope results. In patients with markedly increased vascular permeability, significant quantities of RISA may be lost from blood vessels during the test, which may produce additional error. Severe edema is an example of such a situation. Nevertheless, even under adverse conditions, RISA (and 51Cr) represents a decided advance over hematocrit values for estimating blood volume.

    Central venous pressure measurement. Central venous pressure (CVP) is frequently used as an estimate of blood volume status. However, CVP is affected by cardiac function and by vascular resistance as well as by blood volume. Circumstances in which the CVP does not accurately reflect the relationship between blood quantity and vascular capacity include pulmonary hypertension (emphysema, embolization, mitral stenosis), left ventricular failure, and technical artifacts due to defects in catheter placement and maintenance.

  • Red Blood Cell Substitutes

    Attempts have been made to find an RBC substitute that will not require crossmatching, can be stored conveniently for long periods of time, can be excreted or metabolized in a reasonable period of time, is relatively nontoxic, and can provide an adequate delivery of oxygen to body tissues and return carbon dioxide to the lungs. Thus far, no perfect answer has emerged. The current two leading candidates have been hemoglobin solutions (free of RBC stroma) and synthetic substances, of which the most promising to date are fluorocarbon compounds. However, major problems still remain. Free hemoglobin can precipitate in the tubules of the kidney or alter renal function. Another difficulty involves a generalized and a coronary artery vasoconstrictor effect. Also, free Hb can interfere with some biochemical tests. Fluorocarbons usually must be oxygenated for maximum effectiveness, most commonly by having the patient breathe 100% oxygen. Elimination of fluorocarbons from the body is fairly rapid (the half-life is about 24 hours), which sometimes would necessitate continued administration. Thus far, none of these blood substitute preparations has proved entirely successful. However, several new preparations are now in clinical trials.

  • Albumin and Purified Plasma Protein Fraction

    As mentioned in the earlier discussion about plasma, 5% albumin can be used instead of plasma to restore colloid oncotic pressure, mainly in hypovolemic shock due to massive acute blood loss or extensive burns. About 40% of body albumin is intravascular, with the remainder being in extracellular fluid. In a normal-sized person, 500 ml of blood contains about 11 gm of albumin, which is about 3.5% of total body albumin and about 70% of the albumin synthesized daily by the liver. Therefore, the albumin lost in three or four units of whole blood would be replaced in about 3 days of normal production. The AABB and other investigators believe that albumin has been overused in bleeding persons. They discourage use of albumin infusions in persons with hypoalbuminemia due to chronic liver disease or albumin loss through the kidneys or gastrointestinal tract on the grounds that such therapy does not alter the underlying disease and has only a very short-term effect. They are also critical of therapeutic albumin in hypoalbuminemia due to nutritional deficiency, which should be treated with parenteral hyperalimentation or other nutritional therapy. Purified plasma protein fraction (PPPF) can be used in most cases instead of albumin but has few advantages. It is not recommended when rapid infusion or large PPPF volumes are needed since it may have a hypotensive effect under these conditions. Albumin and PPPF do not transmit viral hepatitis because they are pasteurized.