Hemolytic anemia Hemolytic anemia HA is a complex and serologically HA is a complex and serologically challenging subject, requiring challenging subject, requiring specialized knowledge and lab specialized knowledge and lab experience. experience. The information provided is intended The information provided is intended only as an introduction to the only as an introduction to the subject. When treatment options are subject. When treatment options are listed, they are for information only listed, they are for information only and in no way suggest to clinicians and in no way suggest to clinicians that such options should be taken. that such options should be taken.
Transcript
Hemolytic anemiaHemolytic anemia
HA is a complex and serologically HA is a complex and serologically challenging subject, requiring challenging subject, requiring specialized knowledge and lab specialized knowledge and lab experience.experience.
The information provided is intended The information provided is intended only as an introduction to the subject. only as an introduction to the subject. When treatment options are listed, they When treatment options are listed, they are for information only and in no way are for information only and in no way suggest to clinicians that such options suggest to clinicians that such options should be taken.should be taken.
Hemolytic AnemiaHemolytic Anemia Increased destruction of red blood cellIncreased destruction of red blood cell
s in the peripheral blood without evides in the peripheral blood without evidence of ineffective erythropoiesis is knonce of ineffective erythropoiesis is known as hemolytic anemia. Such anemiawn as hemolytic anemia. Such anemias are generally classified into either ins are generally classified into either inherited or acquired types.herited or acquired types.
The term hemolysis refers to red cell destructiThe term hemolysis refers to red cell destruction in general and should not be interpreted as on in general and should not be interpreted as referring to intravascular red cell destruction oreferring to intravascular red cell destruction only. nly.
It may mean that red cells are removed from tIt may mean that red cells are removed from the bloodstream and hemolysed extravascularlhe bloodstream and hemolysed extravascularly.y.
HA may be defined as primary or secondary diHA may be defined as primary or secondary disease.sease.
There may be no identifiable cause of the aneThere may be no identifiable cause of the anemia, in which case the disease is also termed mia, in which case the disease is also termed “idiopathic”.“idiopathic”.
Hemolysis溶血
HemolysisHemolysis
HemolysisHemolysis is the premature destruction is the premature destruction of RBCs due to intrinsic inherited of RBCs due to intrinsic inherited defects in the RBCs or because of acquiredefects in the RBCs or because of acquired d intravascular abnormalities.intravascular abnormalities.
RBCs normally live about 120 days followRBCs normally live about 120 days following bone marrow release as reticulocytes.ing bone marrow release as reticulocytes. When the RBCs become senescent they When the RBCs become senescent they are removed from the peripheral blood bare removed from the peripheral blood by macrophages in the spleen and liver. y macrophages in the spleen and liver.
Hemolysis may be either intraHemolysis may be either intravascular or extravascularvascular or extravascular
In In intravascular intravascular hemolysishemolysis RBCs lyse i RBCs lyse in the circulation releasing hemoglobin n the circulation releasing hemoglobin into the plasma. Causes include mechainto the plasma. Causes include mechanical trauma, complement fixation, annical trauma, complement fixation, and other toxic damage to the RBC. The fd other toxic damage to the RBC. The fragmented RBCs are calledragmented RBCs are called schistocyte schistocytes s
In In extravascularextravascular hemolysishemolysis RBCs are RBCs are phagocytized by macrophages in the sphagocytized by macrophages in the spleen and liver. Causes include RBC mpleen and liver. Causes include RBC membrane abnormalities such as bound embrane abnormalities such as bound immunoglobulin, or physical abnormaimmunoglobulin, or physical abnormalities restricting RBC deformability thalities restricting RBC deformability that prevent egress from the spleen. Extrat prevent egress from the spleen. Extravascular hemolysis is characterized by vascular hemolysis is characterized by spherocytes.spherocytes.
Intravascular hemolysisIntravascular hemolysis releases hemoglobin releases hemoglobin which is immediately bound by haptoglobin.which is immediately bound by haptoglobin.
Hemoglobin-haptoglobin is cleared almost immHemoglobin-haptoglobin is cleared almost immediately from the plasma by hepatic reticuloendediately from the plasma by hepatic reticuloendothelial cells.othelial cells.
As As intravascular hemolysisintravascular hemolysis with binding to ha with binding to haptoglobin generally overwhelms the rate of happtoglobin generally overwhelms the rate of haptoglobin synthesis, haptoglobin levels decrease.toglobin synthesis, haptoglobin levels decrease.
After haptoglobin is saturated, excess hemogloAfter haptoglobin is saturated, excess hemoglobin is filtered in the kidney and reabsorbed in tbin is filtered in the kidney and reabsorbed in the proximal tubules where the iron is recoverehe proximal tubules where the iron is recovered and converted into ferritin or hemosiderin. d and converted into ferritin or hemosiderin.
Hemoglobinuria indicates severe intravHemoglobinuria indicates severe intravascular hemolysis overwhelming the absascular hemolysis overwhelming the absorptive capacity of the renal tubular cellorptive capacity of the renal tubular cells.s.
Urine hemosiderin is another indicator tUrine hemosiderin is another indicator that intravascular free hemoglobin is beihat intravascular free hemoglobin is being filtered by the kidneys.ng filtered by the kidneys.
Lactic dehydrogenase (LDH) is greatly elLactic dehydrogenase (LDH) is greatly elevated in patients with intravascular evated in patients with intravascular hehemolysismolysis..
Note:Note: Haptoglobin, synthesized by the li Haptoglobin, synthesized by the liver, is decreased in patients with hepatover, is decreased in patients with hepatocelIular disease. celIular disease.
In In extravascular hemolysisextravascular hemolysis spleen an spleen and liver macrophage Fc receptors bind id liver macrophage Fc receptors bind immunoglobulin attached to RBCs and mmunoglobulin attached to RBCs and then either ingest small portions of the then either ingest small portions of the RBC membrane creating spherocytes oRBC membrane creating spherocytes or phagocytizing the RBCs.r phagocytizing the RBCs.
Amino acids from the globin chains arAmino acids from the globin chains are recycled and the Fe removed from the recycled and the Fe removed from the heme and reused.e heme and reused.
The heme is degraded into the tetrapyrThe heme is degraded into the tetrapyrrole, bilirubin. role, bilirubin.
Note: Note: Because little hemoglobin escapBecause little hemoglobin escapes into the plasma in es into the plasma in extravascularextravascular hehemolysismolysis, haptoglobin does not general, haptoglobin does not generally decrease.ly decrease.
Extravascular hemolysisExtravascular hemolysis
Free unconjugated bilirubin is transported to thFree unconjugated bilirubin is transported to the liver where it is conjugated to glucuronic acid.e liver where it is conjugated to glucuronic acid.
In In extravascularextravascular hemolysishemolysis plasma levels of un plasma levels of unconjugated bilirubin increase because the hepatconjugated bilirubin increase because the hepatocytes cannot process the excess bilirubin.ocytes cannot process the excess bilirubin.
Conjugated bilirubin is excreted into the gastroiConjugated bilirubin is excreted into the gastrointestinal tract where it is converted to urobilinontestinal tract where it is converted to urobilinogen and eventually excreted in the feces as stercgen and eventually excreted in the feces as stercobilinogen. obilinogen.
Conjugated bilirubin levels are usually normal iConjugated bilirubin levels are usually normal in n hemolysishemolysis..
3.3. 由抗原特异性淋巴细胞引起：由抗原特异性淋巴细胞引起： AA 。淋巴细胞改变。淋巴细胞改变了识别能力；了识别能力； BB 。耐受性。耐受性 TT 细胞的消失，细胞的消失， BB 细胞细胞得到大力辅助产生自身抗体；得到大力辅助产生自身抗体； CC 。 。 TsTs 细胞功能异细胞功能异常，功能低下，使常，功能低下，使 ThTh 细胞活性相对增高，而使细胞活性相对增高，而使 BB细胞多克隆激活，产生大量自身抗体。细胞多克隆激活，产生大量自身抗体。
4.4. AIHA is often secondary to an existing conditAIHA is often secondary to an existing condition such as lymphoma, SLE, viral infection or ion such as lymphoma, SLE, viral infection or untreated syphilis.untreated syphilis.
AIHA fall into three major AIHA fall into three major categoriescategories
Warm autoantibodies: majority of caseWarm autoantibodies: majority of casess
Cold autoantibodies: minority of casesCold autoantibodies: minority of cases Drug induced hemolytic anemia: respoDrug induced hemolytic anemia: respo
nsible for a small percentage of cases. nsible for a small percentage of cases.
Characteristics of Characteristics of antibodies capable of antibodies capable of
causing hemolytic anemiacausing hemolytic anemia The autoantibody has an optimum reaction The autoantibody has an optimum reaction
temperature of +37℃.temperature of +37℃. The immunoglobulin class is usually IgG, aThe immunoglobulin class is usually IgG, a
nd subclasses IgG3 or IgG1.nd subclasses IgG3 or IgG1. The larger the amount of cell bound antiboThe larger the amount of cell bound antibo
dy, the greater the chance of red cell elimindy, the greater the chance of red cell elimination.ation.
Cold autoantibodies may not react at + 37℃,Cold autoantibodies may not react at + 37℃, but when they lead to complement fixation, but when they lead to complement fixation, red cells are more likely to be eliminated. red cells are more likely to be eliminated.
Warm antibodies WAIHAWarm antibodies WAIHA 温温抗体抗体
Warm autoantibodies are the most coWarm autoantibodies are the most common cause of hemolytic anemia of ammon cause of hemolytic anemia of an immune nature.n immune nature.
The disease may be primary and idiopThe disease may be primary and idiopathic (of no identifable cause), or secoathic (of no identifable cause), or secondary to some underlying condition sundary to some underlying condition such as lymphoma or SLE, or secondary tch as lymphoma or SLE, or secondary to treatment with some medicinal drugo treatment with some medicinal drugs.s.
Lab findings in WAIHALab findings in WAIHA Classically, the Classically, the DAT is strongly positiveDAT is strongly positive. Ho. Ho
wever, when the DAT is positive, it does not alwever, when the DAT is positive, it does not always mean that red cells will be eliminated.ways mean that red cells will be eliminated.
Antibodies are usually IgG and may be complAntibodies are usually IgG and may be complement-binding. ement-binding.
Most antibodies react at + 37℃ by enzyme anMost antibodies react at + 37℃ by enzyme and IAT methods. d IAT methods.
When the autoantibody has strong affinity for When the autoantibody has strong affinity for the antigen, there may be only weak evidence the antigen, there may be only weak evidence of antibody in the serum. This is because wheof antibody in the serum. This is because when cells are strongly auto-sensitized, there may n cells are strongly auto-sensitized, there may be little or no free antibody.be little or no free antibody.
Underlying alloantibodies may becomUnderlying alloantibodies may become detectable following removal of autoe detectable following removal of autoantibodies from patient’s serum by aantibodies from patient’s serum by adsorption of the autoantibody with the dsorption of the autoantibody with the patient’s own cells.patient’s own cells.
Antibody elusions may be Antibody elusions may be carried out with the carried out with the
following resultsfollowing results The antibody detected in an eluate may bThe antibody detected in an eluate may b
e stronger than that in the serum/plasma e stronger than that in the serum/plasma caused by concentration of antibody as a caused by concentration of antibody as a result of the elution procedure. result of the elution procedure.
If the DAT was strongly positive and therIf the DAT was strongly positive and there is no evidence that antibodies were elute is no evidence that antibodies were eluted, an alternative elution technique shoued, an alternative elution technique should be considered, or there is a possibility tld be considered, or there is a possibility that the AIHA is drug induced.hat the AIHA is drug induced.
Cold autoantibodiesCold autoantibodies Cold antibodies of a benign Cold antibodies of a benign
(harmless) nature are frequently (harmless) nature are frequently encountered in blood specimens that encountered in blood specimens that have been cooled. have been cooled.
On rare occasions, a cold On rare occasions, a cold autoantibody may have clinical autoantibody may have clinical significance, and cause hemolytic significance, and cause hemolytic anemia. The disease may be primary anemia. The disease may be primary and idiopathic or secondary to some and idiopathic or secondary to some underlying condition. underlying condition.
Lab findings in cold autoantibLab findings in cold autoantibodiesodies
It may prove impossible to perform a It may prove impossible to perform a DAT or any other red cell tests on a DAT or any other red cell tests on a specimen which becomes agglutinated specimen which becomes agglutinated as it cools.as it cools.
Specimen taken from patients having a Specimen taken from patients having a cold type hemolytic anemia should then cold type hemolytic anemia should then be maintained at +37℃ and kept at be maintained at +37℃ and kept at this temperature for the duration of all this temperature for the duration of all testing, in order to prevent adsorption testing, in order to prevent adsorption of antibody and complement of antibody and complement in vitroin vitro..
Red cells used for the DAT should be wRed cells used for the DAT should be washed in +37℃ saline. ashed in +37℃ saline.
The DAT may be performed on blood cThe DAT may be performed on blood collected into EDTA, which prevents thollected into EDTA, which prevents the e in vitroin vitro uptake of complement. uptake of complement.
Causative antibodies usually have extrCausative antibodies usually have extremely high titers and are IgM. They soemely high titers and are IgM. They sometimes cause hemolysis metimes cause hemolysis in vitroin vitro..
All compatibility tests should be perforAll compatibility tests should be performed at +37℃. The main danger of tranmed at +37℃. The main danger of transfusion is that potent cold agglutinins sfusion is that potent cold agglutinins may mask other clinically significant amay mask other clinically significant alloantibodies and that as a result, allo- lloantibodies and that as a result, allo- as well as auto-incompatible blood is tras well as auto-incompatible blood is transfused.ansfused.
Specific categories of cold autoSpecific categories of cold autoantibodiesantibodies
Cold hemagglutinin diesease (CHAD)Cold hemagglutinin diesease (CHAD) Acute CHAD is secondary to disease (sAcute CHAD is secondary to disease (s
uch as lymphoma) or infection (such auch as lymphoma) or infection (such as s MycoplasmaMycoplasma pneumoniaepneumoniae or infectio or infectious mononucleosis, i.e. glandular fever).us mononucleosis, i.e. glandular fever).Chronic CHAD is usually found in ageChronic CHAD is usually found in aged patients, and sometimes linked to disd patients, and sometimes linked to diseases such as lymphoma. eases such as lymphoma.
Characteristics of CHADCharacteristics of CHAD Children are rarely affected----patients arChildren are rarely affected----patients ar
e usually elderly.e usually elderly. The condition may be self-limiting and flThe condition may be self-limiting and fl
are up only in cold conditions, when the are up only in cold conditions, when the patient’ extremities (ears, nose, fingers patient’ extremities (ears, nose, fingers and toes) become and toes) become cold.cold.
When it occurs with febrile illness, haemWhen it occurs with febrile illness, haemolysis can be increased, but usually remaolysis can be increased, but usually remains extravascular. ins extravascular.
In chronic disease, the autoantibody is In chronic disease, the autoantibody is usually a monoclonal IgM (but sometiusually a monoclonal IgM (but sometimes IgG or IgA) that agglutinates red cmes IgG or IgA) that agglutinates red cells in the cold.ells in the cold.
When the nature of the antibody is bipWhen the nature of the antibody is biphasic (reacts at two different temperathasic (reacts at two different temperatures, in two different ways, i.e. agglutiures, in two different ways, i.e. agglutination at cold temperatures, and haemnation at cold temperatures, and haemolysis when the temperature is raised) olysis when the temperature is raised) it usually leads to a more severe conditit usually leads to a more severe condition.ion.
Complement fixation leads to extravasComplement fixation leads to extravascular red cell destruction and because cular red cell destruction and because of the consumption of complement, mof the consumption of complement, may transiently limit further haemolysis.ay transiently limit further haemolysis.
Complement-binding antibody dissociComplement-binding antibody dissociates ates in vivoin vivo from red cells when they w from red cells when they warm up in the central circulation. The arm up in the central circulation. The antibody then becomes available again,antibody then becomes available again, to rebind with unsensitized red cells to rebind with unsensitized red cells iin vivon vivo..
Lab findings in CHADLab findings in CHAD CHAD is caused by IgM antibody, usuallCHAD is caused by IgM antibody, usuall
y with Anti-I specificity, so it reacts with y with Anti-I specificity, so it reacts with all adult red cells. all adult red cells. Sometimes the causative antibody is antSometimes the causative antibody is anti-I (which may be identified using cord i-I (which may be identified using cord red cells which are I negative, i positive red cells which are I negative, i positive and react more strongly with anti-i).and react more strongly with anti-i).
The antibody may react at +37℃ in the The antibody may react at +37℃ in the presence of albumin.presence of albumin.
The DAT is positive using an antiglobuThe DAT is positive using an antiglobulin reagent specific for complement (alin reagent specific for complement (anti-C3).nti-C3).
The causative antibody is usually of hiThe causative antibody is usually of high titer, agglutinating red cells up to a gh titer, agglutinating red cells up to a titer of 1000 or higher at +4℃.titer of 1000 or higher at +4℃.
Clinical manifestation of Clinical manifestation of CHADCHAD
AnemiaAnemia Reynaud’s syndrome (discolouration of Reynaud’s syndrome (discolouration of
the fingers and toes)the fingers and toes)Treatment options:Treatment options: Transfusion is seldom needed.Transfusion is seldom needed. If transfusion is required, blood should If transfusion is required, blood should
be warmed and washed cells are sometibe warmed and washed cells are sometimes preferred.mes preferred.
Keeping the patient warm at all times is Keeping the patient warm at all times is important.important.
阵发性冷血红蛋白尿阵发性冷血红蛋白尿 PCH is a rare conditioPCH is a rare condition caused by a biphasic (active at two diffen caused by a biphasic (active at two different temperatures) auto anti-P. The antibrent temperatures) auto anti-P. The antibody binds to red cells when the individual ody binds to red cells when the individual becomes cold during the night. As a result becomes cold during the night. As a result of antibody binding, complement also becof antibody binding, complement also becomes cell bound. When the blood warms omes cell bound. When the blood warms as the temperature rises in the morning, tas the temperature rises in the morning, the coated red cells are hemolysed. Hemohe coated red cells are hemolysed. Hemoglobinuria follows, and occurs in episodes globinuria follows, and occurs in episodes related to exposure to cold temperatures. related to exposure to cold temperatures.
Characteristics of PCHCharacteristics of PCH PCH usually occurs secondary to infection iPCH usually occurs secondary to infection i
n children, when antibodies are produced thn children, when antibodies are produced that cross-react with P antigen. at cross-react with P antigen.
PCH may be seen post PCH may be seen post M. pneumoniaeM. pneumoniae infec infection (or other infections such as measles, mtion (or other infections such as measles, mumps, chicken pox, CMV, Epstein Barr virus umps, chicken pox, CMV, Epstein Barr virus and syphilis).and syphilis).
Sufferers are highly susceptible when enviroSufferers are highly susceptible when environmental temperature is low.nmental temperature is low.
Disease is usually transient and self-limiting,Disease is usually transient and self-limiting, especially in children. especially in children.
Lab findings in PCHLab findings in PCH The cause is usually a polyclonal IgG antibody, The cause is usually a polyclonal IgG antibody,
that reacts as a biphasic hemolysin using the that reacts as a biphasic hemolysin using the Donath-Landsteiner test (this is carried out at Donath-Landsteiner test (this is carried out at cold and then warm temperatures to demonstrcold and then warm temperatures to demonstrate the biphasic nature of the antibody; red celate the biphasic nature of the antibody; red cells are sensitized at +4℃, and bind complement ls are sensitized at +4℃, and bind complement which hemolyses the red cells when the test is which hemolyses the red cells when the test is warmed to +37℃). warmed to +37℃). The Donath-Landsteiner test is not carried out The Donath-Landsteiner test is not carried out in blood transfusion service labs, but could be in blood transfusion service labs, but could be performed in a pathology lab with a hematoloperformed in a pathology lab with a hematology department.gy department.
Blood grouping and compatibility testiBlood grouping and compatibility testing is complicated by autoantibodies. ng is complicated by autoantibodies.
IgG anti-P is usually the responsible anIgG anti-P is usually the responsible antibody.tibody.
DAT is positive using antiglobulin reagDAT is positive using antiglobulin reagent with anti-C3 specificity only.ent with anti-C3 specificity only.
Clinical manifestation in Clinical manifestation in PCHPCH
HemoglobinuriaHemoglobinuria Anemia-usually mild but may be severeAnemia-usually mild but may be severe
Treatment options:Treatment options: Transfusion may be needed, using the lTransfusion may be needed, using the l
east incompatible red cells.east incompatible red cells. Corticosteroids may be given to treat seCorticosteroids may be given to treat se
vere anemia.vere anemia. Patients should avoid exposure to cold Patients should avoid exposure to cold
temperature. temperature.
Drug induced hemolytic Drug induced hemolytic anemiaanemia
药物诱发的溶血性贫血药物诱发的溶血性贫血 DIHA has the poteDIHA has the potential to be extremely serious and may be ntial to be extremely serious and may be life-threatening. Many theories have beelife-threatening. Many theories have been postulated on how drugs induce hemon postulated on how drugs induce hemolytic anemia. lytic anemia. Certain medicinal drugs-either prescribCertain medicinal drugs-either prescribed or over-the-counter, may have the sided or over-the-counter, may have the side effect of triggering AIHA. This may alse effect of triggering AIHA. This may also apply to certain chemnical and insectio apply to certain chemnical and insecticides. cides.
In drug induced hemolytic anemia, the In drug induced hemolytic anemia, the DAT is usually positive using an anti-IgDAT is usually positive using an anti-IgG antiglobulin reagent. The sensitizing G antiglobulin reagent. The sensitizing antibody is usually IgG with Rh specificantibody is usually IgG with Rh specificity.ity.
Individuals with glucose-6-phosphate dIndividuals with glucose-6-phosphate dehydrogenase (G6PD) deficency may exehydrogenase (G6PD) deficency may experience a hemolytic episode if given drperience a hemolytic episode if given drugs containing aspirin. Anti-malaria drugs containing aspirin. Anti-malaria drugs are also a potential cause of acute hugs are also a potential cause of acute hemolysis. So the drug is the trigger for aemolysis. So the drug is the trigger for an intrinsic deficiency.n intrinsic deficiency.
Process of drug Process of drug autoimmune hemolytic autoimmune hemolytic
anemiaanemiaDrugs can induce AIHA in various ways, Drugs can induce AIHA in various ways,
including the following:including the following: Antibody to the drug: drugs may be Antibody to the drug: drugs may be
immunogenic in themselves and immunogenic in themselves and antibodies formed against them cross-antibodies formed against them cross-react with the patient’s red cells.react with the patient’s red cells.
Antibody to the red cell membrane: drugs Antibody to the red cell membrane: drugs may have the potential to bind loosely to may have the potential to bind loosely to red cells, initiating an immune response red cells, initiating an immune response that then auto-sensitizes the red cells.that then auto-sensitizes the red cells.
Antibody to the drug-membrane compAntibody to the drug-membrane complex: a single specificity antibody may blex: a single specificity antibody may be formed against the immunogen create formed against the immunogen created by cell bound drug + red cell membed by cell bound drug + red cell membrane together. rane together.
Drug metabolites: the antibody may be Drug metabolites: the antibody may be formed against a chemical produced bformed against a chemical produced by the drug rather than the drug itself. y the drug rather than the drug itself.
Lab findings in drug related Lab findings in drug related AIHAAIHA
The DAT may be The DAT may be strongly positivestrongly positive with with or without complement involvement.or without complement involvement.
If an eluate is prepared from the patieIf an eluate is prepared from the patient’s cells, the eluate will not react witnt’s cells, the eluate will not react with routine reagent cells but only with ceh routine reagent cells but only with cells coated with the specific drug. lls coated with the specific drug.
Routine antibody screening tests may Routine antibody screening tests may be negative. be negative.
Some drug induced antibodies show blood Some drug induced antibodies show blood group specificity (e.g. Rh and Duffy)group specificity (e.g. Rh and Duffy)
Penicillin-induced hemolytic anemia:Penicillin-induced hemolytic anemia:-positive DAT with or without cell destruct-positive DAT with or without cell destructionion-penicillin-bound red cells are sensitized b-penicillin-bound red cells are sensitized by the autoantibodyy the autoantibody
Cephalosporin-induced hemolytic anemia:Cephalosporin-induced hemolytic anemia:-this drug is related to penicillin and has a -this drug is related to penicillin and has a similar pattern as above.similar pattern as above.-cephalosporin-induced HA is becoming -cephalosporin-induced HA is becoming more prevalent. more prevalent.
Treatment options for drug Treatment options for drug induce AIHAinduce AIHA
If the clinical condition persists If the clinical condition persists unnoticed and drugs continue to be unnoticed and drugs continue to be administered, the situation may become administered, the situation may become debilitating and could lead to fatalities. debilitating and could lead to fatalities.
When the AIHA is the result of the When the AIHA is the result of the production of anti-drug antibodies, production of anti-drug antibodies, discontinuation of the drug leads to discontinuation of the drug leads to eventual resolution of the hemolytic eventual resolution of the hemolytic anemia and the DAT becomes negative anemia and the DAT becomes negative quite rapidly.quite rapidly.
If the drug associated AIHA is the resulIf the drug associated AIHA is the result of the production of red cell autoantit of the production of red cell autoantibodies, removal of the drug may not hbodies, removal of the drug may not halt the autoimmune process. Various dalt the autoimmune process. Various drugs may cause this, e.g. rugs may cause this, e.g. αα -methyldop-methyldopa.a.
Red cell Red cell abnormalities/deficienciesabnormalities/deficienciesConditions caused by the malfunction of inheriConditions caused by the malfunction of inherited genes can result in red cell membrane defited genes can result in red cell membrane deficiencies, or in enzyme or hemoglobin malfuncciencies, or in enzyme or hemoglobin malfunctions. tions. For example: thalassemia is a hemoglobin defiFor example: thalassemia is a hemoglobin deficiency in which red cells have a shortened lifesciency in which red cells have a shortened lifespan. Sickle cell anemia is caused by structural pan. Sickle cell anemia is caused by structural abnormal within the hemoglobin molecule. abnormal within the hemoglobin molecule. Red cells with abnormal shapes, such as the spRed cells with abnormal shapes, such as the spherical cells seen in hereditary spherocytosis, aherical cells seen in hereditary spherocytosis, are also prone to hemolysis. Red cells from patire also prone to hemolysis. Red cells from patients with paroxysmal nocturnal hemoglobinurients with paroxysmal nocturnal hemoglobinuria (PNH) are susceptible to hemolysis because a (PNH) are susceptible to hemolysis because of a red cell membrane defect. of a red cell membrane defect.
This is an acquired intrinsic defect of This is an acquired intrinsic defect of the red cell membrane.the red cell membrane.
The cells have an increased The cells have an increased sensitivity to traces of complement sensitivity to traces of complement and become hemolytic readily.and become hemolytic readily.
Patients with bone marrow failure Patients with bone marrow failure may develop PNH.may develop PNH.
Characteristics of PNHCharacteristics of PNH
Ongoing hemolysis is more commonly Ongoing hemolysis is more commonly seen than hemolytic episodes.seen than hemolytic episodes.
PNH can be life-threatening.PNH can be life-threatening.
Lab findings in PNHLab findings in PNH
Red cells may lyse in a low ionic strengtRed cells may lyse in a low ionic strength saline medium.h saline medium.
The Ham’s acid hemolysis test is positiThe Ham’s acid hemolysis test is positive. In the Ham’s acid test, red cells are ve. In the Ham’s acid test, red cells are tested for their resistance to lysis in an atested for their resistance to lysis in an acidified serum; lowering the pH causes ccidified serum; lowering the pH causes complement lysis in PNH. (This test is not omplement lysis in PNH. (This test is not carried out in blood transfusion service lcarried out in blood transfusion service labs, but could be performed in a patholoabs, but could be performed in a pathology lab with a hematology department).gy lab with a hematology department).
Clinical manifestation in Clinical manifestation in PNHPNH
Treatment options for Treatment options for patients with PNHpatients with PNH
Transfusion may modulate the producTransfusion may modulate the production of PNH cells by the bone marrow.tion of PNH cells by the bone marrow.
PNH responds to corticosteroids.PNH responds to corticosteroids. Prophylactic use of anticoagulants (sucProphylactic use of anticoagulants (suc
h as Warfarin) may be beneficial.h as Warfarin) may be beneficial. In very severe forms of PNH, bone maIn very severe forms of PNH, bone ma
rrow transplantation may be considererrow transplantation may be considered. d.
THE DIRECT ANTIGLOBULIN TEST
The DAT should be performed as part of a differential diagnosis on every patient in whom the presence of hemolysis has been established.
The predictive value of a positive DAT is 83% in a patient with hemolytic anemia, but only 1.4% in a patient without hemolytic anemia.
Small amounts of IgG and complement appear to be present on all red cells.
Healthy individuals can have 5 to 90 IgG molecules/red cell and 5 to 40 C3d molecules/red cell; these levels are below the threshold of detection in routine testing.
Depending on the technique and reagents used, the DAT can detect 100 to 500 molecules of IgG/red cell and 400 to 1100 molecules of C3d/red cell.
Positive DATs are reported in 1:1000 up to 1:14,000 blood donors and 1% to 15% of hospital patients.
Most blood donors with positive DATs appear to be perfectly healthy and most patients with positive DATs have no obvious signs of hemolytic anemia although, after careful evaluation, some may show slight evidence of increased red cell destruction.
Although a positive DAT in a patient with hemolytic anemia indicates the most likely diagnosis is one of the immune hemolytic anemias, the DAT can be positive, coincidentally, in patients with hemolytic anemia that is not immune-mediated.
Conversely, some patients with immune hemolytic anemia have a negative DAT . .
Positive DATs due to elevated levels of IgG or complement, with no clear correlation with anemia, have been noted on the red cells of patients with sickle cell disease, β-thalasemia, renal disease, multiple myeloma, auto-immune disorders, AIDS, and other diseases with elevated serum globulin or blood urea nitrogen (BUN) levels.
Interpretation of positive DATs should include the patient’s history, clinical data, and the results of other laboratory tests.
When investigating a transfusion reaction, performance of the DAT on postreaction specimens is part of the initial transfusion reaction investigation.
The DAT may be positive if sensitized red cells have not been destroyed or negative if hemolysis and rapid clearance have occurred.
Evaluation of a Positive DAT
A positive DAT alone is not diagnostic. The interpretation of the significance of this
positive result requires knowledge of the patient’s diagnosis; recent drug, pregnancy, and transfusion history; and information on the presence of acquired or unexplained hemolytic anemia.
Dialogue with the attending physician is important. Clinical considerations together with laboratory data should dictate the extent to which a positive DAT is evaluated.
Patient History The following situations may warrant further investi
gation of a positive DAT. 1. History of recent transfusion.
When a patient has recently been transfused, a positive DAT may be the first indication of a developing immune response. The developing antibody sensitizes the transfused red cells that have the corresponding antigen and the DAT becomes positive; the antibody may not be of sufficient quantity to be detected in the serum. Antibody may appear as early as 7 to 10 days after transfusion in primary immunization or as early as 1 to 2 days in a secondary response; these alloantibodies could shorten the survival of red cells already transfused or given in subsequent transfusions. A mixed-field appearance in the posttransfusion DAT (ie, agglutination of donor red cells and no agglutination of the patient’s red cells), may or may not be observed.
2. Administration of drugs previously associated with immune-mediated hemolysis. Many drugs have been reported to cause a positive DAT and/or immune-mediated hemolysis, but the occurrence is rare.
3. History of hematopoietic progenitor cell or organ transplantation. Passenger lymphocytes of donor origin produce antibodies directed against ABO or other blood group antigens on the recipient’s cells, causing a positive DAT.
4. Administration of IVIG or IV anti-D. Intravenous immunoglobulin (IVIG) may contain ABO antibodies, anti-D, or, sometimes, other antibodies. Intravenous Rh Immune Globulin used to treat immune thrombocytopenic purpura causes Rh-positive patients to develop a positive DAT.
If an anemic patient with a positive DAT does show evidence of hemolysis, testing to evaluate a possible immune etiology is appropriate.
Reticulocytosis, spherocytes observed on the peripheral blood film, hemoglobinemia, hemoglobinuria, decreased serum haptoglobin, and elevated levels of serum unconjugated bilirubin or lactate dehydrogenase (LDH), especially LDH1, may be associated with increased red cell destruction.
If there is no evidence of immune hemolysis, no further studies are necessary, unless the patient requires red cell transfusion and the serum contains incompletely identified unexpected antibodies to red cell antigens; an eluate may be helpful with antibody identification
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