Diagnosis of hemolytic anemia in adults

Author: Wilma Barcellini, MD

Section Editor: Robert A Brodsky, MD
Deputy Editor: Jennifer S Tirnauer, MD

Contributor Disclosures

All topics are updated as new evidence becomes available and our peer review process is complete.

Literature review current through: Feb 2024. | This topic last updated: Jul 08, 2022.

INTRODUCTION

Hemolytic anemia is defined as anemia due to a shortened survival of circulating red

blood cells (RBCs) due to their premature destruction. There are numerous causes of
hemolytic anemia, including inherited and acquired conditions, acute and chronic
processes, and mild to potentially life-threatening severity. Occasionally the cause
will be obvious from the history, physical examination, or findings on the peripheral
blood smear, but often the ultimate diagnosis requires a synthesis of all of this
information and additional laboratory testing.

The key clue that suggests that hemolysis is the cause of the anemia is an increase in
the reticulocyte count that is not explained by recent bleeding or recent correction of
iron or other nutrient deficiency, or other causes (pregnancy, acclimatization to
altitude). Reticulocytosis should be expressed as an absolute number rather than a
percentage, and it should be related to the hemoglobin and hematocrit values.
Patients may also have evidence of RBC destruction including increased lactate
dehydrogenase (LDH) and unconjugated bilirubin, decreased haptoglobin, and RBC
shape changes on the peripheral blood smear.

The causes of hemolytic anemia and a diagnostic approach to the adult with
unexplained hemolytic anemia are discussed here. Other topic reviews present
general approaches to determining the cause of anemia and diagnosis of specific
types of hemolytic anemia:

General approaches:
● General approach, child – (See "Approach to the child with anemia".)
● General approach, adult – (See "Diagnostic approach to anemia in adults".)

Specific types of hemolytic anemia:

● Immune-mediated:

• Warm autoimmune hemolytic anemia (AIHA), child – (See "Autoimmune

hemolytic anemia (AIHA) in children: Classification, clinical features, and
diagnosis".)
• Warm AIHA, adult – (See "Warm autoimmune hemolytic anemia (AIHA) in
adults".)
• Paroxysmal cold hemoglobinuria (PCH) – (See "Paroxysmal cold
hemoglobinuria".)
• Cold agglutinin disease (CAD) – (See "Cold agglutinin disease".)
• Drug-induced hemolytic anemia – (See "Drug-induced hemolytic anemia".)
● Heritable/genetic:

• Hemoglobinopathies – (See "Hemoglobinopathy: Screening and counseling in

the reproductive setting and fetal diagnosis" and "Diagnosis of thalassemia
(adults and children)" and "Diagnosis of sickle cell disorders".)
• RBC membrane abnormalities – (See "Hereditary spherocytosis" and
"Hereditary elliptocytosis and related disorders" and "Hereditary
stomatocytosis (HSt) and hereditary xerocytosis (HX)".)
• RBC enzyme disorders – (See "Diagnosis and management of glucose-6-
phosphate dehydrogenase (G6PD) deficiency" and "Pyruvate kinase
deficiency" and "Rare RBC enzyme disorders".)
• Congenital dyserythropoietic anemia (CDA) – (See "Overview of causes of
anemia in children due to decreased red blood cell production", section on
'Congenital dyserythropoietic anemia'.)
● Other/less common:

• Microangiopathic hemolytic anemia (MAHA) such as thrombotic

thrombocytopenic purpura (TTP), hemolytic uremic syndrome (HUS), or drug-
induced thrombotic microangiopathy (DITMA) – (See "Diagnostic approach to
suspected TTP, HUS, or other thrombotic microangiopathy (TMA)".)
• Infectious and traumatic causes – (See "Non-immune (Coombs-negative)
hemolytic anemias in adults".)
• Hemolysis associated with blood transfusion – (See "Hemolytic transfusion
reactions" and "Approach to the patient with a suspected acute transfusion
reaction".)
 • Paroxysmal nocturnal hemoglobinuria (PNH) – (See "Clinical manifestations
and diagnosis of paroxysmal nocturnal hemoglobinuria".)

CONCEPTUAL FRAMEWORK

Causes of hemolysis can be categorized in various ways, including whether the

abnormality is intrinsic or extrinsic to the red blood cell (RBC) (intracorpuscular
versus extracorpuscular defects), whether the condition is inherited or acquired,
whether the hemolysis is acute or chronic, whether the mechanism involves immune
destruction due to antibodies (immune versus non-immune mechanism), and
whether the hemolysis occurs in the vasculature or in the reticuloendothelial
macrophages in the liver and spleen (intravascular versus extravascular hemolysis).
Most of the inherited conditions are intracorpuscular, and most of the hemolysis by
an immune mechanism is extravascular.

RBC turnover — The typical lifespan of a red blood cell (RBC) is approximately 110 to
120 days (four months). During this time, RBCs are subject to remarkable mechanical
stresses as they traverse capillaries and splenic cords much smaller than their
diameter, which requires repeated cycles of deformation and elastic recoil. This
happens many millions of times over the course of the RBC lifespan.

The properties of the RBC that allow it to withstand this stress include a highly
deformable membrane and underlying cytoskeleton, an optimal surface-to-volume
ratio, and an enzymatic system that continually restores the proper redox
environment of the cell. The optimal membrane surface is approximately 40 percent
greater than that of a perfectly spherical cell of equivalent volume. RBC volume is
regulated by ion pumps and channels that control the entry of water and cations
including sodium, potassium, calcium, and magnesium. Metabolic enzymes generate
ATP needed by the cation pumps; 2,3-BPG, which regulates oxygen uptake and
release by hemoglobin; and reducing capacity (eg, glutathione, NADPH), to protect
the oxygen-rich RBC from oxidant injury [1].

Normal aging of RBCs results in age-dependent RBC destruction. This typically

occurs at a rate of approximately 1 percent of RBCs daily. The percentage of RBCs
that are cleared from the circulation can be increased dramatically in hemolytic
states, especially when an enlarged spleen is contributing to hemolysis [2]. (See "Red
blood cell survival: Normal values and measurement".)
Hemolysis occurs when the RBC is unable to maintain its intact structure during
passages through the circulation and reticuloendothelial system. Hemolysis by any
mechanism stimulates a compensatory increase in RBC production via increased
erythropoietin (EPO) secretion by the kidney. EPO in turn stimulates the bone
marrow to produce more RBC precursors, which is followed within a few days by an
increase in the reticulocyte count and within an additional one to two days by an
increase in the hemoglobin level and hematocrit. (See "Regulation of erythropoiesis"
and "Diagnostic approach to anemia in adults", section on 'Reticulocyte production'.)

The occurrence of anemia and its severity is determined by the balance between the
extent of hemolysis and the capacity of the bone marrow to amplify RBC production.
This balance is illustrated in the following calculations and clinical examples:
● RBC survival and turnover rate can be calculated under steady state conditions
(eg, when the secretion and response to EPO are intact) from the percentage of
reticulocytes and the reticulocyte lifespan (RLS). The RLS increases proportionally
as the hematocrit decreases and reticulocytes enter the circulation at
progressively earlier stages in their maturation (when the hematocrit is lower,
reticulocytes are released earlier from the bone marrow). In the steady state
without anemia the typical RLS is approximately one day, after which the cell
becomes a mature RBC. In severe anemia, reticulocytes can be released from
the bone marrow as much as 1.5 days early ("shift cells"), giving a RLS of 2.5
days.

• RBC survival – RBC survival (days) ≈ 100 ÷ [Reticulocytes (percent) ÷ RLS (days)]

The RLS is 1.0, 1.5, 2.0, or 2.5 days at hematocrits of 45, 35, 25, and 15
percent, respectively ( figure 1). For an individual without hemolysis who has
a hematocrit of 40 and a reticulocyte count of 1 percent, RBC survival is
calculated to be approximately 100 days (100 ÷ [1 ÷ 1] = 100). Corrections for
the increased RLS can be made in patients with severe anemia to reflect the
degree of reticulocytosis more accurately. (See 'High reticulocyte count'
below.)

• RBC turnover – The rate of RBC turnover is the reciprocal of RBC survival:

RBC turnover rate (percent/day) = 100 ÷ RBC survival (days)

In adults, the normal rate of RBC turnover is approximately 1 percent per day,
and the maximal sustainable capacity of the bone marrow to increase RBC
 production in an adult is approximately 5 percent per day (ie, approximately
five times normal). In children, the bone marrow capacity can increase RBC
production up to eight times normal.

Reticulocytosis (increase in the production of new RBCs, evidenced by appearance of

reticulocytes in the peripheral blood) requires adequate iron and vitamins (B12,
folate) for RBC production, along with a normally functioning bone marrow and
adequate erythropoietin production. Reticulocytosis may not occur in individuals
who are deficient in iron, vitamin B12, or folate; those with concomitant bone
marrow abnormalities (from infection or primary bone marrow disorders); and those
with inadequate erythropoietin production (from chronic kidney disease). (See
'Hemolysis without reticulocytosis' below.)

There are several free applications that calculate the reticulocyte index from the
reticulocyte percentage and hematocrit, allowing identification of inadequate bone
marrow compensation.

Additionally, a bone marrow responsiveness index (BMRI) has been calculated as

([absolute reticulocyte count] × [patient Hb/normal Hb]) to discriminate an anemia
with effective erythropoiesis from those with ineffective erythropoiesis, such as in
congenital dyserythropoietic anemia. The BMRI has been extended to other
hemolytic conditions including autoimmune hemolytic anemia (AIHA) and hereditary
spherocytosis [3-5]. (See 'High reticulocyte count' below.)

Additional information about the pathogenesis of specific types of hemolytic anemia

are discussed in separate topic reviews. (See "Hereditary spherocytosis", section on
'Pathophysiology' and "Pathogenesis of paroxysmal nocturnal hemoglobinuria" and
"Pathophysiology of thalassemia" and "Cold agglutinin disease", section on
'Pathogenesis' and "Warm autoimmune hemolytic anemia (AIHA) in adults", section
on 'Pathogenesis'.)

Intracorpuscular versus extracorpuscular causes of hemolysis — Classifying

hemolytic anemias according to whether the abnormality resides within the RBC
itself (intracorpuscular) versus external to the RBC (extracorpuscular) is helpful
because it incorporates the patient history; it also allows the clinician to determine
whether the hemolysis is reversible, whether transfused RBCs will also be affected,
and whether the treatment should be directed at a specific RBC defect or at another
condition (eg, an infection or drug).
The table lists the causes of hemolytic anemia classified by whether the underlying
cause is intracorpuscular or extracorpuscular ( table 1).
● Intracorpuscular causes include abnormalities of hemoglobin structure and
function, membrane structure and function, and cytoplasmic composition
including metabolic mechanisms controlling RBC volume and redox potential.
(See 'Intracorpuscular' below.)
● Extracorpuscular causes are those in which external factors lead to premature
loss of membrane, membrane structural damage, volume gain or loss, changes
in the solubility of hemoglobin, and changes in the redox state of cellular
proteins. (See 'Extracorpuscular' below.)

Intracorpuscular — Intrinsic (intracorpuscular) RBC causes are those in which the

altered properties of the RBC are responsible for hemolysis. These defects include
the following three major categories [6,7]:
● Hemoglobinopathies – Hemoglobinopathies include sickle cell disease (SCD),
thalassemia, and unstable hemoglobin variants. These affect the solubility of
hemoglobin. When hemoglobin becomes insoluble, it can precipitate and
damage the RBC membrane. Some abnormal hemoglobins are less able to
recover from oxidant challenge, leading to the formation of Heinz bodies
(denatured hemoglobin), as in Hgb Köln. (See "Diagnosis of sickle cell disorders"
and "Diagnosis of thalassemia (adults and children)" and "Unstable hemoglobin
variants".)
● RBC membrane/cytoskeletal disorders – Disorders that affect the structure of
the RBC membrane and underlying cytoskeleton include hereditary
spherocytosis (HS), hereditary elliptocytosis (HE), and hereditary stomatocytosis
(HSt). These conditions may be associated with a suboptimal membrane surface-
area-to-volume ratio (eg, HS) or a defect in a membrane protein with ion channel
function such as Band 3 that may alter salt and water handling, leading to
altered surface-membrane-to-volume ratio. (See "Hereditary spherocytosis" and
"Hereditary elliptocytosis and related disorders" and "Hereditary stomatocytosis
(HSt) and hereditary xerocytosis (HX)".)

End-stage liver disease or cirrhosis can lead spur cell anemia with severe
hemolysis resulting from altered lipid metabolism. The membrane changes
render erythrocytes less flexible and prone to hemolysis. Spur cell anemia due to
 end-stage liver disease has a poor prognosis and is often fatal within a few
weeks if liver transplantation is not performed.
● RBC metabolic abnormalities – Metabolic abnormalities include deficiency of
glucose-6-phosphate dehydrogenase (G6PD) and pyruvate kinase (PK). These
defects affect the metabolic capacity of the RBC, which promotes solute
transport and recovery from oxidant damage. (See "Diagnosis and management
of glucose-6-phosphate dehydrogenase (G6PD) deficiency" and "Pyruvate kinase
deficiency" and "Hereditary stomatocytosis (HSt) and hereditary xerocytosis
(HX)".)

The vast majority of the intracorpuscular RBC causes are inherited, and conversely,
the majority of the inherited disorders are intracorpuscular. Exceptions include rare
conditions such as paroxysmal nocturnal hemoglobinuria (PNH), an acquired
intracorpuscular abnormality caused by expansion of a clone of RBCs that are
hypersensitive to complement lysis; acquired alpha thalassemia in the setting of
myelodysplasia, an acquired intracorpuscular abnormality caused by expansion of a
clone of RBCs that harbor a genetic variant in the gene that encodes alpha globin;
and hereditary TTP, an inherited condition in which abnormalities in the
microvasculature lead to episodes of microangiopathic hemolysis. (See "Clinical
manifestations and diagnosis of paroxysmal nocturnal hemoglobinuria" and "Clinical
manifestations, diagnosis, and classification of myelodysplastic syndromes (MDS)"
and "Hereditary thrombotic thrombocytopenic purpura (hTTP)".)

Extracorpuscular — Extrinsic (extracorpuscular) causes are those in which the RBCs

are normal but are destroyed due to mechanical, immunologic, infectious, or
metabolic/oxidant damage. These abnormalities are almost always acquired. The
major extracorpuscular causes of hemolysis include:
● Antibody-mediated – Antibodies directed against RBC membrane components
(eg, autoimmune hemolytic anemia [AIHA], alloimmune hemolytic anemia, acute
hemolytic transfusion reaction [AHTR], delayed hemolytic transfusion reaction
[DHTR], some drug-induced hemolytic anemias). (See "Warm autoimmune
hemolytic anemia (AIHA) in adults", section on 'Pathogenesis' and "Cold
agglutinin disease", section on 'Pathogenesis' and "Immunologic transfusion
reactions" and "Drug-induced hemolytic anemia".)

Less common antibody-mediated disorders include mixed AIHA (with

concomitant warm and cold autoantibodies) and paroxysmal cold
 hemoglobinuria (PCH). (See "Paroxysmal cold hemoglobinuria" and "Warm
autoimmune hemolytic anemia (AIHA) in adults".)

Administration of Rho(D) immune globulin to an RhD-positive individual (eg, for

treatment of immune thrombocytopenia) or administration of intravenous
immune globulin (IVIG) can also promote antibody-mediated RBC destruction.
(See "Initial treatment of immune thrombocytopenia (ITP) in adults", section on
'Anti-D as an alternative to IVIG' and "Intravenous immune globulin: Adverse
effects", section on 'Hemolysis'.)

Severe forms of AIHA are associated with solid organ transplant and
hematopoietic stem cell transplantation.
● Drug induced hemolytic anemia – Several drugs may induce hemolytic
anemia, including historically well-described drugs and newer drugs such as
immune checkpoint inhibitors. (See "Drug-induced hemolytic anemia".)
● Paroxysmal nocturnal hemoglobinuria (PNH) – PNH is caused by a
somatically-acquired variant in the phosphatidylinositol glycan anchor
biosynthesis, class A (PIGA) gene, resulting in a deficiency of
glycosylphosphatidyl-inositol-anchored proteins (GPI-AP), including complement
regulatory proteins CD55 and CD59. The disease is characterized by chronic
intravascular hemolysis, increased susceptibility to infections, bone marrow
failure, and deep vein thrombosis (DVT). (See "Pathogenesis of paroxysmal
nocturnal hemoglobinuria" and "Clinical manifestations and diagnosis of
paroxysmal nocturnal hemoglobinuria".)
● Hypersplenism – Stasis, trapping, and destruction of RBC in an enlarged spleen
(hypersplenism). The spleen is also the major site of removal of warm antibody-
coated red cells and most of the intracorpuscular defects. (See "Splenomegaly
and other splenic disorders in adults", section on 'Hypersplenism'.)
● Mechanical trauma – Mechanical trauma to the RBCs secondary to high velocity
jets (malfunctioning cardiac valves, ventricular assist devices); fibrin stands
across vessels that shear RBCs in disseminated intravascular coagulation (DIC);
or platelet microthrombi in TTP, hemolytic uremic syndrome (HUS), or drug-
induced thrombotic microangiopathy (DITMA). (See "Non-immune (Coombs-
negative) hemolytic anemias in adults", section on 'Fragmentation'.)
 ● Oxidant exposure – Exposure to compounds with oxidant potential (eg, aniline
dyes, dapsone, phenazopyridine) in individuals with an underlying metabolic
defect such as G6PD deficiency, congenital methemoglobinemia, or unstable
Hgb variants (eg, sulfonamides), as well as those without an underlying defect.
(See "Diagnosis and management of glucose-6-phosphate dehydrogenase
(G6PD) deficiency" and "Unstable hemoglobin variants" and
"Methemoglobinemia", section on 'Acquired causes'.)
● Infectious diseases – Destruction of RBC by pathogens such as malaria,
babesiosis, bartonellosis, or clostridium perfringens ( picture 1). (See "Non-
immune (Coombs-negative) hemolytic anemias in adults".)
● Toxins and poisons – Less common causes include snake and insect bites,
certain toxins, thermal burns, and copper poisoning (eg, Wilson disease).
Although rare, Wilson disease should be considered in every child and young
adult presenting with hemolytic anemia, since this complication can be fatal [8].
(See "Drug-induced hemolytic anemia".)

Immune versus non-immune — Immune hemolysis generally refers to RBC

destruction by antibodies and/or complement proteins bound to the RBC surface.
Immune hemolysis is characterized by a positive direct antiglobulin test (DAT; also
called direct Coombs test) and/or a positive indirect antiglobulin test (also called
indirect Coombs test, or, in the setting of pretransfusion testing, referred to as an
antibody screen).

Site of RBC destruction — The site of hemolysis can be intravascular (within the
circulation) or extravascular (via the reticuloendothelial macrophages and
monocytes of the liver, spleen, bone marrow, and lymph nodes). The site often
determines the clinical severity and immediate management needs of the patient.
This is because the site of RBC destruction determines whether the byproducts of
RBC destruction such as free hemoglobin will be routed through the
reticuloendothelial system or will be liberated directly into the circulation.
● If products of hemolysis are liberated into the circulation, they will appear as
free serum hemoglobin or heme and urinary hemoglobin, heme, or
hemosiderin. The serum and urine may be pink or darker brown. (See
'Diagnostic approach' below.)
 ● Free hemoglobin or heme in the circulation can cause significant damage to the
kidney, causing acute renal failure, and can trigger DIC or increase the risk of
thrombosis [9,10]. (See 'Immediate management issues before the cause is
identified' below and 'Thrombotic complications' below.)

The site of RBC destruction is one of the main determinants of clinical severity. As an
example, RBC destruction in AIHA is generally extravascular, as reticuloendothelial
macrophages progressively phagocytize small pieces of RBC membrane that are
opsonized with autoantibodies. In contrast, in hemolysis from mechanical trauma
(eg, prosthetic heart valve, marching, bongo drums), the lysis can be immediate and
complete within the circulation, before the RBCs reach the reticuloendothelial
system. Likewise, in conditions in which the membrane attack complex (MAC) of
complement creates a hole in the RBC membrane (eg, paroxysmal nocturnal
hemoglobinuria [PNH], paroxysmal cold hemoglobinuria [PCH], and severe forms of
cold agglutinin disease) a large component of intravascular hemolysis is likely.
Generally, extravascular hemolysis is less severe than intravascular one.

However, in many cases there is a component of both intravascular and

extravascular hemolysis, especially when hemolysis is severe. The site of RBC
destruction thus is helpful in determining the cause of hemolysis and the need for
more aggressive therapy but typically is not diagnostic of a specific condition.

Intravascular hemolysis
● Definition and clinical findings – Intravascular hemolysis refers to hemolysis
that occurs primarily within the vasculature. This occurs when there is a
considerable amount of structural damage to the RBC membrane (eg,
mechanical shearing, complement MAC) or when the reticuloendothelial system
becomes overwhelmed [11]. When severe, intravascular hemolysis is
characterized by pink or brown serum and dark urine with free serum and urine
hemoglobin [12]. The pink color is due to oxyhemoglobin and the brownish color
is due to the oxidized form, methemoglobin.

Free hemoglobin binds to haptoglobin, and the hemoglobin-haptoglobin

complex is rapidly removed by the liver, leading to a reduction in plasma
haptoglobin, often to undetectable levels (see 'High LDH and bilirubin; low
haptoglobin' below). Dimers of alpha-beta globin that are not bound by
haptoglobin are small enough (molecular weight 34,000 daltons) to be filtered
by the glomerulus and appear in the urine as hemoglobinuria.
 Urine hemosiderin may be seen several days after an episode of intravascular
hemolysis, as renal tubular cells take up the heme, degrade it, store it as
hemosiderin, and eventually are shed into the urine. Urine hemosiderin is
detected using Prussian blue staining (iron stain) of the urine sediment. (See
'Immediate management issues before the cause is identified' below.)
● Causes – Causes of intravascular hemolysis include the following ( table 1):

• Direct trauma, as in bongo drummers and march hemoglobinuria (runners' or

foot-strike hemolysis)
• Shear stress, as in defective mechanical heart valves
• Heat damage, as in thermal burns
• Complement-induced lysis, as in paroxysmal nocturnal hemoglobinuria (PNH)
• Osmotic lysis following infusion of hypotonic solutions
• Lysis from bacterial toxins (eg, clostridial sepsis)
• Lysis from exposure to high concentrations of copper
• Thrombotic microangiopathies (TMA) including TTP, HUS, or drug-induced
TMA
• Acute hemolytic transfusion reaction
• Administration of Rho(D) immune globulin to RhD-positive individuals, such as
for treatment of immune thrombocytopenia (ITP)
• Immune hemolysis that overwhelms the reticuloendothelial system
Extravascular hemolysis — Extravascular hemolysis refers to hemolysis that
occurs primarily via macrophages of the reticuloendothelial system in the liver,
spleen, bone marrow, and lymph nodes. Severely damaged RBCs, especially those
coated with complement, are primarily destroyed in the liver, an organ that receives
a larger proportion of the cardiac output than the spleen. Poorly deformable RBCs
such as spherocytes or sickled cells are primarily destroyed in the spleen, in the
cords of Billroth. These unique vascular channels end blindly, unlike other vascular
channels in the body. The only way for a RBC with a diameter of 7 to 8 microns to
escape from these cords and return to the general circulation is to deform
sufficiently to pass through 2 to 3 micron slits in the walls of the cords ( picture 2).
Senescent or damaged RBCs remain in the cords and are phagocytosed by
monocytes and macrophages.

RBCs destroyed in the spleen are usually phagocytosed in their entirety and digested
within phagosomes of macrophages. Most of the hemoglobin is degraded to release
heme, with each molecule of heme converted to equimolar amounts of biliverdin,
iron, and carbon monoxide via the action of microsomal heme oxygenase [13-16].
● The biliverdin is immediately reduced to unconjugated bilirubin by the enzyme
biliverdin reductase and is released into the plasma. (See "Bilirubin
metabolism".)
● Under normal circumstances, the iron is efficiently released from macrophages
into the plasma, mediated by the iron export protein ferroportin. It is
transported to the bone marrow and used in the production of new RBCs. In
patients with the anemia of chronic disease/inflammation, iron release is
impaired due to the action of hepcidin, resulting in impaired iron mobilization
for new RBC production. (See "Regulation of iron balance" and "Anemia of
chronic disease/anemia of inflammation", section on 'Hepcidin (primary
regulator of iron homeostasis)'.)
● The carbon monoxide formed from heme breakdown initially binds to the
hemoglobin of intact RBCs as carboxyhemoglobin. It is subsequently released in
the pulmonary capillaries and excreted into the expired air.

LIST OF CAUSES

The major causes of hemolytic anemia include:

● Autoimmune

• Warm autoimmune hemolytic anemia (AIHA)

• Cold agglutinin disease (CAD)
• Paroxysmal cold hemoglobinuria (PCH)
● Congenital hemolytic anemias
• Alpha thalassemia
• Beta thalassemia
• Glucose-6-phosphate dehydrogenase (G6PD) deficiency
• Hereditary spherocytosis (HS)
• Hereditary elliptocytosis (HE)
• Hereditary stomatocytosis (HSt)
• Hereditary xerocytosis (HX)
• Pyruvate kinase (PK) deficiency
 • Sickle cell disease
• Unstable hemoglobin variants
● Disseminated intravascular coagulation (DIC)
● Drug-induced hemolytic anemias
• Drug-induced immune hemolysis
• Drug-induced hemolysis associated with G6PD deficiency
• Drug-induced thrombotic microangiopathy (DITMA)
● Transfusion-related hemolysis
• Acute hemolytic transfusion reaction (AHTR)
• Delayed hemolytic transfusion reaction (DHTR)
● Other conditions
• Clostridial sepsis
• Mechanical hemolysis from aortic stenosis or prosthetic heart valve
• Mechanical hemolysis from marching or bongo drumming
• Osmotic lysis from hypotonic infusion
• Paroxysmal nocturnal hemoglobinuria (PNH) receiving anti-complement
therapy
• RBC parasite (eg, malaria, Babesia)
• Snake bite
• Thrombotic microangiopathy (TMA) such as thrombotic thrombocytopenic
purpura (TTP) or hemolytic uremic syndrome (HUS)
• Spur cell anemia from end-stage liver disease
These can be classified in a number of ways, including mechanism (intracorpuscular
versus extracorpuscular; immune versus non-immune), site of hemolysis
(intravascular versus extravascular), and acuity/duration (acute versus chronic). (See
'Conceptual framework' above.)

DIAGNOSTIC APPROACH

Overview of the evaluation — Some patients with especially severe anemia

(hemoglobin <6 g/dL) or other worrisome findings may require immediate life-saving
interventions before hemolysis has been diagnosed and/or the cause identified [17].
Importantly, life-saving interventions such as transfusion for severe anemia;
plasmapheresis for possible thrombotic thrombocytopenic purpura (TTP); or
vigorous hydration and diuresis for an acute transfusion reaction should not be
withheld while confirming the diagnosis. (See 'Immediate management issues
before the cause is identified' below.)

Recognizing and diagnosing hemolytic anemia in patients with a classic presentation

is straightforward. However, for many patients, the exact timing of the onset of
anemia may be unclear, the red blood cell (RBC) morphology may be unrevealing, or
there may be several possible diagnoses under consideration. Other patients may
have more than one cause of anemia, one of which may blunt the normal
reticulocyte response to hemolysis.

The diagnosis of hemolytic anemia is suspected in a patient with chronic or new

onset symptoms of anemia (eg, fatigue, weakness, shortness of breath), a low
hemoglobin level, and an increased reticulocyte count that is not explained by
accelerated RBC production due to recent bleeding; repletion of iron, vitamin B12,
folate, or copper; or administration of erythropoietin. Reticulocytosis (expressed as
absolute number) may be absent or inadequate to Hb levels in conditions with lack
of bone marrow compensation.

Additional laboratory testing is often used to confirm the diagnosis of hemolytic

anemia and to determine the likely cause. Testing to confirm hemolysis may be done
simultaneously (increased unconjugated bilirubin and lactate dehydrogenase [LDH],
and decreased haptoglobin) with testing to determine the cause, or sequentially.
(See 'Laboratory confirmation of hemolysis' below and 'Post-diagnostic testing to
determine the cause' below.)

Once the diagnosis of hemolytic anemia is relatively certain, our approach to

determining the cause and providing immediate interventions is as follows
( algorithm 1):
● Rapidly identify and triage individuals with potentially life-threatening conditions
that require urgent involvement of a specialist (often a hematologist or
transfusion medicine physician), such as a thrombotic microangiopathies (TMA),
disseminated intravascular coagulation (DIC), or an acute transfusion reaction.
● Obtain a thorough history and physical examination targeted to potential causes
of hemolysis.
● If the patient has a clear history of life-long anemia or classic findings on the
peripheral blood smear that suggest one of the hereditary hemolytic anemias
 due to hemoglobinopathy, metabolic defect, or membrane defect, pursue
specific testing for the likely condition.
● For other patients, obtain a direct antiglobulin (Coombs) test (DAT) to determine
whether the anemia is immune or non-immune.
● If the DAT is positive, immune hemolysis is the likely diagnosis. Some patients
may require additional testing to identify associated conditions.
● If the DAT is negative, perform specific diagnostic tests suggested by the patient
history and physical examination. These may be obtained sequentially or
simultaneously depending on the urgency of the evaluation.
● Other laboratory testing that reveals one of the specific causes of hemolysis is
also strongly supportive and in many cases sufficient for diagnosis (see 'Post-
diagnostic testing to determine the cause' below); in some cases this may be
available at the time of the initial evaluation and in others it may be obtained as
part of the evaluation.

Hematologic consultation should be obtained in virtually all patients with a new

onset of hemolysis, since sudden and life-threatening worsening of anemia may
occur, requiring urgent coordination between treating clinicians, clinical
pathologists, and transfusion medicine or blood bank personnel for appropriate
management. Hemolysis may also be the first sign of an underlying systemic
disorder (eg, thrombotic thrombocytopenic purpura, systemic lupus erythematosus,
chronic lymphocytic leukemia) and may require an urgent intervention to prevent
death or disease-related complications.

History and physical examination — A systematic approach, starting with a

thorough history and physical examination is the cornerstone of the evaluation.
Helpful clues from the history and physical examination include the following, if
present:
● Rapid onset of symptoms of anemia in the absence of bleeding is consistent
with brisk hemolysis.
● Jaundice is consistent with brisk hemolysis that overwhelms the capacity of the
reticuloendothelial system to convert heme to storage iron. Mild jaundice is also
present in chronic hemolysis.
● Dark urine and highly increased LDH is consistent with intravascular hemolysis.
 ● Recent blood transfusion suggests possible acute hemolytic transfusion
reaction; transfusion in the previous four weeks also raises the possibility of a
delayed hemolytic transfusion reaction.
● Initiation of a new medication with potential for causing hemolysis suggests
possible drug-induced etiology.
● History of hemolytic anemia or unexplained anemia in family members suggests
an inherited disorder; this is more likely if multiple first degree family members
are affected.
● History of pigmented gallstones or presence of gallstones implies chronic
hemolysis that overwhelms the reticuloendothelial system.
● Splenomegaly suggests expansion of the reticuloendothelial capacity.

However, the absence of these features does not eliminate the possibility of
hemolytic anemia. Patients with chronic compensated hemolytic anemia may have
minimal to no symptoms of anemia, a negative family history, no new drugs, and no
evidence of jaundice or splenomegaly.

Laboratory confirmation of hemolysis — There is no single specific diagnostic test

for hemolytic anemia. However, most experts consider the diagnosis to be accepted
if most of the following findings are present:
● Anemia that is not due to another obvious cause.
● Increased reticulocyte count that is not explained by accelerated RBC production
due to recent bleeding; repletion of iron, vitamin B12, folate, or copper; or
administration of erythropoietin.
● Signs of RBC destruction such as increased lactate dehydrogenase (LDH), low
haptoglobin, increased unconjugated bilirubin.

Additional test results that are consistent with a specific cause of hemolytic anemia
(schistocytes or spherocytes on peripheral blood smear; free hemoglobin or pink
serum; newly positive direct antiglobulin [Coombs] test [DAT]; hemoglobin analysis
demonstrating an abnormal hemoglobin) are highly supportive and in some cases
diagnostic if present, but their absence does not exclude the possibility of hemolysis.
(See 'Post-diagnostic testing to determine the cause' below.)

CBC/blood smear review — All patients for whom the diagnosis of hemolytic
anemia is considered will have had a complete blood count. In the majority of cases,
this will include a white blood cell (WBC) count, platelet count, and RBC indices.
Hemolysis will be associated with some degree of anemia in the majority of cases,
especially when it first occurs, when there is a delay in the production of new RBCs to
compensate for hemolysis, and, if it is severe, when the bone marrow cannot fully
compensate. (See 'RBC turnover' above.)

Review of the peripheral blood smear is an extremely valuable tool for determining
the presence and cause of hemolytic anemia. In many cases, findings on the blood
smear are essential to providing life-saving treatment. Examples include:
● Thrombotic microangiopathies (TMAs) such as thrombotic thrombocytopenic
purpura (TTP) or drug-induced TMA (DITMA) have microangiopathic hemolysis
with schistocytes.
● Infections such as malaria or Babesia have visible microorganisms on a thick
smear.
● Bite cells suggest hemolysis due to an oxidant drug in a patient with glucose-6-
phosphate dehydrogenase (G6PD) deficiency.
● Microspherocytes suggest warm autoimmune hemolytic anemia (warm AIHA) or
drug-induced hemolytic anemia.
● Spherocytes, elliptocytes, and stomatocytes suggest hereditary RBC membrane
disorders.
● RBC agglutination suggests cold-agglutinin disease.
● Abundant spur cells in conjunction with severe liver disease suggest spur cell
anemia.

An initial evaluation for the cause of anemia may have already occurred in some
individuals for whom the likelihood of hemolysis was thought to be low or for whom
hemolysis was not considered initially. The discussion herein presumes that other
common causes of anemia have been evaluated or excluded based on the clinical
history, examination, or laboratory testing. (See "Diagnostic approach to anemia in
adults".)

High reticulocyte count — A high reticulocyte count implies an accelerated

production of RBCs in the bone marrow. Increased reticulocytes is a typical finding in
hemolytic anemia but is not specific for hemolysis; the bone marrow can also
increase RBC production in response to bleeding, nutrient repletion (eg, vitamin B12,
folic acid, iron), or erythropoietin administration. The absence of reticulocytosis does
not eliminate the possibility of hemolysis, as some individuals have concomitant
bone marrow suppression or reduced bone marrow function that interferes with
production of reticulocytes. (See 'Hemolysis without reticulocytosis' below.)

The degree of reticulocytosis can be estimated from the peripheral blood smear, as
reticulocytes are larger than mature RBCs, lack central pallor, and have a bluish tint
(polychromasia) ( picture 3). The count can be quantified from a manual count on a
peripheral blood smear stained for reticulin ( picture 4) or by an automated
counter. Many counters automatically provide the reticulocyte count. (See
"Automated complete blood count (CBC)", section on 'Reticulocytes'.)

Reticulocytes can be expressed as a percentage of RBCs or as an absolute number;

these numbers can be corrected for the degree of anemia and the lifespan of the
reticulocytes in the circulation. In cases of markedly increased reticulocytosis or
failure of the bone marrow to produce reticulocytes, any of these measures is likely
to be a relatively good indicator of the degree of reticulocytosis. However, the
corrections make the count more accurate, and in some cases are essential to
determining whether reticulocytosis is truly increased:
● Reticulocyte percentage – The reticulocyte percentage conveys the percentage
of all RBCs that are reticulocytes. Since it is relative to the total RBC count, it may
be artificially increased in severe anemia and artificially decreased if the patient
is not anemic. The normal reticulocyte percentage in a patient without hemolysis
is in the range of 1 to 2 percent. In patients with hemolysis and an otherwise
intact bone marrow, reticulocyte percentage is at least 4 to 5 percent, and often
considerably higher. This was illustrated in case series such as the following:

• A series of 109 individuals with autoimmune hemolytic anemia, in which the

median reticulocyte percentage at diagnosis was 9 percent [18]. However, the
range was large (0.4 to 92 percent) and approximately one-fifth had a
reticulocyte percentage <4 percent.

• A subsequent series of 308 patients with AIHA found that reticulocytopenia

was associated with a severe anemia (Hb <6 g/dL), indicating an inadequate
bone marrow compensation that possibly contributed to the clinical severity
[4,5].
● Corrected reticulocyte count – The reticulocyte percentage is a relative
number. For any given number of reticulocytes, a lower the total number of
 RBCs (the denominator) will raise the percentage of these RBCs that are
reticulocytes. Thus, the reticulocyte percentage can be multiplied by the
patient's hematocrit divided by a normal hematocrit (eg, 45 percent) to give a
corrected reticulocyte percentage. As an example, if reticulocytes are 10 percent
in a patient with a hematocrit of 22.5 percent, the corrected count can be
calculated as follows:

10 percent x (22.5 percent ÷ 45 percent) = 10 percent x 0.5 = 5 percent.

● Absolute reticulocyte count – The absolute reticulocyte count has the

advantage of accurately reflecting the degree of reticulocytosis regardless of the
degree of anemia and the number of RBCs [19,20]. The normal absolute
reticulocyte count is between 25,000 to 75,000/microL (ie, approximately 1
percent of an absolute RBC count of 5,000,000 cells/microL). An example of the
utility of this measure is a severely anemic patient with a hematocrit of 18
percent, a RBC count of 2,000,000/microL, and reticulocyte count of 3 percent.
While the reticulocyte count of 3 percent appears high, the absolute reticulocyte
count is only 60,000/microL (ie, 3 percent of 2,000,000), which is in the normal
range and does not reflect an adequate bone marrow compensation.
● Corrected absolute reticulocyte count – The more severe the anemia, the
younger the reticulocytes are when they are released into the circulation, and
hence the longer their lifespan in the circulation. The absolute reticulocyte count
can be corrected for the reticulocyte lifespan (RLS), also called the reticulocyte
maturation time (RMT). The RLS is 1.0, 1.5, 2.0, or 2.5 days at hematocrits of 45,
35, 25, and 15 percent, respectively ( figure 1). (See 'RBC turnover' above.)

In a patient with a hematocrit of 18 percent and an absolute reticulocyte count

of 60,000/microL, the RLS is approximately 2.5 days; thus, the corrected absolute
reticulocyte count is (60,000 ÷ 2.5 = 24,000/microL), which is inappropriately low.
● Reticulocyte production index (RPI) – The RPI makes corrections for both the
hematocrit and the reticulocyte lifespan (calculator 1):

RPI = Reticulocytes (percent) x (HCT ÷ 45) x (1 ÷ RMT)

The RPI in an individual without hemolysis or blood loss is approximately 1. A

value in excess of 2 to 3 is considered increased, whereas a value <2 in a patient
with anemia is considered inappropriately low [18].
There are several free applications that calculate the reticulocyte index from the
reticulocyte percentage and hematocrit, allowing the identification of inadequate
bone marrow compensation. A bone marrow responsiveness index (BMRI) has been
calculated as [(absolute reticulocyte count) × (patient Hb/normal Hb)] to discriminate
an anemia with effective erythropoiesis from those with ineffective erythropoiesis,
such as congenital dyserythropoietic anemia (CDA). The BMRI has then been
extended to other hemolytic conditions including AIHA and hereditary spherocytosis
[3-5].

The use of the reticulocyte percentage and absolute reticulocyte count are discussed
in more detail separately. (See "Diagnostic approach to anemia in adults".)

High LDH and bilirubin; low haptoglobin — The other major typical finding in
hemolytic anemia aside from reticulocytosis is evidence of RBC destruction from
breakdown products. Hemolysis releases lactate dehydrogenase (LDH) and
hemoglobin from RBCs. Hemoglobin is bound by circulating haptoglobin, which
facilitates heme recycling; hemoglobin is converted to bilirubin as part of the
degradation and heme recycling process ( figure 2).

Thus, high LDH and bilirubin and low haptoglobin are all consistent with hemolysis
as summarized in the table ( table 2). Caveats include:
● LDH and bilirubin – High LDH and bilirubin are not very specific for hemolysis,
as there are numerous other possible causes of these abnormalities ( table 3).
When bilirubin elevation is due to hemolysis, the elevation is predominantly in
the indirect (unconjugated) bilirubin. (See "Classification and causes of jaundice
or asymptomatic hyperbilirubinemia".)
● Haptoglobin – The normal range for serum haptoglobin is wide. A low
haptoglobin is likely to be due to hemolysis, and an undetectable haptoglobin
level is almost always due to hemolysis. In a series of 100 patients with various
medical conditions, a haptoglobin level of 25 mg/dL provided the best cutoff
between hemolytic and non-hemolytic disorders [21]. The sensitivity and
specificity of a haptoglobin ≤25 mg/dL were 83 and 96 percent. However, a
normal or increased haptoglobin does not eliminate the possibility of hemolysis
because haptoglobin is an acute phase reactant that can be increased in the
setting of inflammation (see "Acute phase reactants"). Other causes of low
haptoglobin include hepatic insufficiency, abdominal trauma, and congenital
ahaptoglobinemia [22].
In one series of reports, the combination of an increased serum LDH and a reduced
haptoglobin was 90 percent specific for diagnosing hemolysis, while the combination
of a normal serum LDH and a serum haptoglobin >25 mg/dL was 92 percent
sensitive for ruling out hemolysis [21,23].

Immediate management issues before the cause is identified — Certain

management issues may need to be addressed before the specific cause of
hemolytic anemia is definitively established. Importantly, life-saving interventions
should not be delayed while awaiting the results of diagnostic testing.
● Rate of hemoglobin decline – It is important to have a sense of the rate of
decline in the hemoglobin level to manage the patient properly. Individuals with
a very slow decline in hemoglobin may be able to adapt to and tolerate severe
anemia without end organ ischemia, whereas those with brisk hemolysis and a
rapid drop in hemoglobin level may be quite symptomatic and require more
aggressive treatment and accelerated evaluation, even if the absolute
hemoglobin level is not that low.
● Transfusion – Patients with severe anemia (hemoglobin <6 g/dL, higher
hemoglobin in some cases with heart or lung disease) should be transfused with
RBCs, especially if there is active bleeding, symptoms of organ ischemia, or
ongoing brisk hemolysis. It may be appropriate to obtain a pre-transfusion
blood sample that can be stored for later analysis, particularly if an inherited
cause of hemolytic anemia is suspected. For patients with possible immune-
mediated hemolysis for whom crossmatch compatible blood cannot be
identified, blood designated for immediate release can be transfused. This and
other alternatives such as RBC genotyping or autoadsorption are discussed in
more detail separately. (See "Pretransfusion testing for red blood cell
transfusion", section on 'Emergency release blood for life-threatening anemia or
bleeding' and "Red blood cell (RBC) transfusion in individuals with serologic
complexity", section on 'Identifying compatible units'.)
● Plasma exchange – In cases of presumed thrombotic microangiopathy (TMA; ie,
microangiopathic hemolytic anemia and thrombocytopenia), diagnostic testing
may take hours to days. The use of therapeutic plasma exchange for a
presumptive diagnosis of thrombotic thrombocytopenic purpura (TTP) and
information regarding the diagnostic evaluation are presented separately. (See
 "Diagnostic approach to suspected TTP, HUS, or other thrombotic
microangiopathy (TMA)".)
● Complement blockade – C5 inhibition with eculizumab or ravulizumab should
be considered in patients with thrombotic microangiopathy (TMA) with
ADAMTS13 activity >10 percent and suspected complement-mediated TMA (CM-
TMA) or paroxysmal nocturnal hemoglobinuria (PNH). (See "Thrombotic
microangiopathies (TMAs) with acute kidney injury (AKI) in adults: CM-TMA and
ST-HUS", section on 'Terminal complement blockade' and "Treatment and
prognosis of paroxysmal nocturnal hemoglobinuria".)

Proximal complement inhibition (anti C1s) with sutimlimab is used in cold

agglutinin disease (CAD). (See "Cold agglutinin disease".)
● Hydration and hemodynamic support – For individuals with apparent severe
intravascular hemolysis (eg, acute hemolytic transfusion reaction [AHTR] due to
ABO incompatible transfusion), free hemoglobin in the circulation can cause
renal failure, hypotension, and disseminated intravascular coagulation.
Aggressive hydration and other supportive measures are discussed separately.
(See "Approach to the patient with a suspected acute transfusion reaction",
section on 'Suspected acute hemolytic reaction' and "Evaluation and
management of disseminated intravascular coagulation (DIC) in adults", section
on 'Treatment' and "Clinical features and diagnosis of heme pigment-induced
acute kidney injury".)

Post-diagnostic testing to determine the cause

Obvious cause- proceed to specific testing — In some of the obvious/classic

presentations of hemolytic anemia, it may make sense to proceed directly to specific
diagnostic testing ( algorithm 1):
● Anemia and thrombocytopenia with numerous schistocytes ( picture 5) on the
blood smear suggests a TMA such as TTP, HUS, or drug-induced TMA. (See
"Diagnostic approach to suspected TTP, HUS, or other thrombotic
microangiopathy (TMA)".)
● Rapid onset of fever, back pain, dark urine, and pink plasma following a blood
transfusion suggests an AHTR. (See "Hemolytic transfusion reactions", section
on 'Acute hemolytic transfusion reactions' and "Approach to the patient with a
 suspected acute transfusion reaction", section on 'Acute hemolytic transfusion
reaction (AHTR)'.)
● Lifelong anemia, splenomegaly, and RBC morphology typical of one of the
inherited disorders such as spherocytes ( picture 6), elliptocytes ( picture 7),
or stomatocytes ( picture 8) suggests a congenital RBC membrane/cytoskeletal
defect. (See "Hereditary spherocytosis" and "Hereditary elliptocytosis and
related disorders" and "Hereditary stomatocytosis (HSt) and hereditary
xerocytosis (HX)".)
● A blood smear characteristic of sickle cell disease ( picture 9) or thalassemia
( picture 10) in a patient with classic findings suggests a hemoglobinopathy.
(See "Diagnosis of sickle cell disorders" and "Diagnosis of thalassemia (adults
and children)".)
● A rapid drop in hemoglobin level after exposure to a drug known to cause
hemolysis suggests drug-induced hemolytic anemia, which may be due to
glucose-6-phosphate dehydrogenase (G6PD) deficiency ( table 4). (See
"Diagnosis and management of glucose-6-phosphate dehydrogenase (G6PD)
deficiency".)
● Hemoglobinuria associated with pancytopenia or acute onset thrombosis
(especially abdominal vein or central venous sinus thrombosis) suggests
paroxysmal nocturnal hemoglobinuria (PNH), which is evaluated with flow
cytometry. (See "Clinical manifestations and diagnosis of paroxysmal nocturnal
hemoglobinuria", section on 'Flow cytometry/FLAER'.)

Cause not obvious - start with Coombs test — In some cases, the cause of
hemolysis may be less clear. As examples, the presence of numerous spherocytes
could indicate autoimmune hemolytic anemia or hereditary spherocytosis.

For such individuals, we use the direct antiglobulin (Coombs) test (DAT) to distinguish
between hemolysis due to an immune mechanism (eg, AIHA) and hemolysis that is
less likely to be due to an immune mechanism ( algorithm 1).

The DAT is used to determine whether patient RBCs are coated with IgG,
complement, or both. The assay is performed by taking washed patient RBCs and
incubating them with anti-human IgG and anti-human C3d antibodies. In AIHA, anti-
human antibodies and/or anti-C3d antibodies form bridges (agglutination) between
red cells by binding the human antibodies on the patient's red cells. Agglutination is
graded visually as negative to 4+, as is illustrated in the figure ( figure 3). Less
frequently, RBC may be coated with IgM or IgA, and their presence is revealed by
specific antisera.

A DAT should be obtained in all patients who present with anemia, laboratory
evidence of hemolysis (ie, increased lactate dehydrogenase, increased
indirect/unconjugated bilirubin, reduced haptoglobin), and an absence of
schistocytes on the peripheral blood smear. It is important to distinguish between
warm-AIHA and cold agglutinin disease. (See "Cold agglutinin disease".)

Coombs test interpretation

● Warm AIHA – Warm AIHA is generally due to an IgG that binds to RBCs at a
temperature around 37°C, and the DAT is typically positive with anti-IgG antisera
or IgG plus complement at low titer.
● Cold agglutinin disease – Cold agglutinin disease is due to an IgM
autoantibody that has an optimum temperature of reaction at 4° C (thermal
range 4 to 34°C) and strongly activates complement. The DAT is positive with
anti-complement antisera and a high titer of cold agglutinins is found in the
serum ( picture 11). Typically, spontaneous agglutination of RBCs occurs at
room temperature, sometimes invalidating automated blood counts and raising
the diagnostic suspect.
● Mixed AIHA – Mixed forms of AIHA show common characteristics of warm and
cold autoantibodies, with a DAT positive for both IgG and complement, along
with high titer cold agglutinins. As an exception, paroxysmal cold
hemoglobinuria (PCH) is due to an IgG that binds to RBCs in the cold but causes
severe intravascular hemolysis at 37°C. PCH is diagnosed with the Donath-
Landsteiner test. (See "Paroxysmal cold hemoglobinuria", section on 'Evaluation
and diagnosis'.)
● Atypical forms – Atypical AIHA forms include:

• IgA driven – IgA-driven cases are due to an IgA antibody, which is frequently
but not always associated with an IgG.

• Warm IgM – AIHAs due to IgM with a thermal range close to body
temperature (warm IgM) are rare and cause extremely serious AIHA that is
potentially fatal. These IgM antibodies are able to strongly activate
complement in vivo and cause massive intravascular hemolysis. They may
 appear weakly positive for complement on the DAT or may be DAT negative,
possibly delaying diagnosis.

• DAT-negative – The DAT can be performed with various methods, the most
classic being the test tube with polyspecific and monospecific antisera (anti-
IgG, anti-complement, anti-IgA, and anti-IgM). More sensitive tests can reveal
smaller amounts of antibodies bound to RBCs; these tests include
microcolumn and solid phase tests; these are widely used in routine
diagnosis. More sophisticated methods involve washing RBCs with low ionic
strength solutions (LISS), improving the ability to detect low affinity
autoantibodies, and experimental tests such as immunoradiometric assays,
ELISA, and cytometry. Despite these methodological improvements, 5 to 10
percent of AIHAs remain DAT-negative, making diagnosis especially
challenging.
● False-positive DAT – The DAT may be falsely positive after the administration of
various immunoglobulin-containing therapies:

• Intravenous immune globulin (IVIG)

• RhD immune globulin
• Antithymocyte globulin
• Daratumumab
The DAT may also be falsely positive in diseases with elevated serum gamma
globulins or paraproteins.

Moreover, the DAT may be positive due to alloantibodies in patients who have
recently been transfused, such as during delayed hemolytic transfusion
reactions (DHTRs) and in the hemolytic disease of the fetus and newborn
(HDFN).

The DAT may be positive without clinical evidence of AIHA in a small percentage
of healthy blood donors (<0.1 percent) and in hospitalized patients (0.3 to 8
percent).

Selected testing to further narrow the diagnosis — Other features that may be
useful to test for in narrowing the diagnostic possibilities include the following:
● Evidence of intravascular hemolysis (eg, pink serum, positive serum free
hemoglobin, positive urine dipstick for heme, positive urine for hemosiderin)
suggests one of the following:
 • AHTR
• Overwhelming bacterial infection (eg, from clostridium perfringens)
• Paroxysmal nocturnal hemoglobinuria (PNH)
• Paroxysmal cold hemoglobinuria (PCH)
Red to brown urine in a patient with a normal plasma color may be due to
transient hemolysis (if the samples were not evaluated simultaneously) or to a
cause other than intravascular hemolysis (eg, myoglobinuria, beet ingestion).
● Splenomegaly suggests a congenital, infectious, or neoplastic process. (See
"Splenomegaly and other splenic disorders in adults".)
● Abnormal finding on blood smear:

• Spherocytes ( picture 6), microspherocytes, and elliptocytes ( picture 7)

suggest AIHA, assessed by DAT; or hereditary spherocytosis, assessed by tests
for reduced eosin-5-maleimide (EMA) binding, increased osmotic fragility,
and/or genetic testing. Elliptocytosis may also suggest myelodysplasia,
assessed by bone marrow evaluation with chromosomal analysis. (See "Warm
autoimmune hemolytic anemia (AIHA) in adults" and "Hereditary
spherocytosis" and "Clinical manifestations, diagnosis, and classification of
myelodysplastic syndromes (MDS)".)

• Acanthocytes (spur cells) ( picture 12) and target cells suggest liver disease.
(See "Non-immune (Coombs-negative) hemolytic anemias in adults", section
on 'Liver and kidney disease'.)

• Blister or "bite" cells ( picture 13) suggest oxidant injury in the setting of
G6PD deficiency. (See "Diagnosis and management of glucose-6-phosphate
dehydrogenase (G6PD) deficiency".)

• Red cell "ghosts" ( picture 1) indicate severe intravascular hemolysis, most

often associated with overwhelming bacterial infection (eg, from clostridium
perfringens).

Involvement of specialists with expertise in hemolytic anemias, laboratory diagnosis,

or genetic testing may be helpful in especially challenging cases.

ATYPICAL PRESENTATIONS
Anemia is often multifactorial, and concomitant disorders that blunt the normal
reticulocyte response can call the diagnosis of hemolytic anemia into question. (See
'Hemolysis without reticulocytosis' below.)

In some individuals, compensation for hemolysis may be sufficient to raise the

hemoglobin into the normal range. (See 'Hemolysis without anemia' below.)

In other cases, an increased reticulocyte count initially thought to indicate hemolysis

may in fact be due to another cause of increased erythropoiesis. (See 'Reticulocytosis
without hemolysis' below.)

Hemolysis without reticulocytosis — Hemolytic anemia can be seen in the absence

of an appropriate reticulocyte response, often resulting in a more profound degree
of anemia [24]. This occurs when the bone marrow is not capable of responding
appropriately to anemia. If hemolysis is suspected or confirmed but the reticulocyte
count is inappropriately low, there are several possible concomitant conditions that
may be responsible for blunting the reticulocyte response:
● Iron deficiency (absolute or functional) – (See "Causes and diagnosis of iron
deficiency and iron deficiency anemia in adults" and "Diagnosis of iron
deficiency in chronic kidney disease" and "Treatment of anemia in nondialysis
chronic kidney disease".)
● Deficiency of vitamin B12, folate, or copper – (See "Clinical manifestations and
diagnosis of vitamin B12 and folate deficiency" and "Sideroblastic anemias:
Diagnosis and management", section on 'Copper deficiency'.)
● Anemia of chronic inflammation (anemia of chronic disease) – (See "Anemia of
chronic disease/anemia of inflammation".)
● Alcohol – (See "Hematologic complications of alcohol use".)
● Myelodysplasia, aplastic anemia, or other primary bone marrow disorder – (See
"Clinical manifestations, diagnosis, and classification of myelodysplastic
syndromes (MDS)" and "Aplastic anemia: Pathogenesis, clinical manifestations,
and diagnosis".)
● Transient red blood cell (RBC) aplasia due to parvovirus infection, which targets
erythropoietic precursor cells [25,26] – (See "Clinical manifestations and
diagnosis of parvovirus B19 infection".)
 ● Drug-induced bone marrow suppression such as therapy for chronic
lymphocytic leukemia (CLL) [27] – (See "Overview of the complications of chronic
lymphocytic leukemia", section on 'Anemia'.)

Other settings in which the reticulocyte response may be inappropriately low include
an autoimmune hemolytic anemia in which the autoantibody also targets RBC
progenitor cells in the bone marrow, or a transient lag in the production of
reticulocytes during the first few days of new-onset hemolysis.

Hemolysis without anemia — Hemolysis without anemia can be seen if the bone
marrow capacity to increase RBC production is sufficient to overcome the anemia
caused by the hemolysis. (See 'RBC turnover' above.)

Despite a normal hemoglobin and hematocrit, hemolysis can still be detected from
increased reticulocyte count, increased serum LDH, and decreased serum
haptoglobin. An estimate of RBC turnover from the reticulocyte count in non-anemic
patients should yield a value of ≤5 or ≤8 percent per day in adults and children,
respectively.

Reticulocytosis without hemolysis — A patient thought to have hemolytic anemia

based on an increased reticulocyte count may in fact have another cause of
reticulocytosis. If the clinical picture is not fully consistent with hemolytic anemia or a
specific cause of hemolysis cannot be identified, it may be appropriate to evaluate
the patient for other causes of reticulocytosis such as:
● Recovery from an episode of bleeding or ongoing bleeding.
● Repletion of iron, vitamin B12, or folate in a patient who was deficient.
● Administration of erythropoietin.
● Recovery from a bone marrow insult such as an infection (eg, parvovirus),
medication, or alcohol.

THROMBOTIC COMPLICATIONS

There is a well-known association between hemolytic anemia and thrombosis, which

may be seen with either intravascular or extravascular hemolysis. The mechanism(s)
are not well understood. Postulated factors include the effects of free plasma heme
or hemoglobin, depletion of nitric oxide (NO), splenectomy, antiphospholipid
antibodies in some patients with autoimmune hemolysis, and prothrombotic
changes in the surface of affected RBCs [28].
Management and prevention of thrombosis in specific disease settings is discussed
in separate topic reviews:
● Paroxysmal nocturnal hemoglobinuria (PNH) – (See "Pathogenesis of paroxysmal
nocturnal hemoglobinuria", section on 'Thrombosis'.)
● Sickle cell disease (SCD) – (See "Overview of the pulmonary complications of
sickle cell disease", section on 'Venous thromboembolism and pulmonary
thrombosis'.)
● Autoimmune hemolytic anemia (AIHA) – (See "Warm autoimmune hemolytic
anemia (AIHA) in adults", section on 'Thromboembolic complications'.)
● Hereditary spherocytosis (HS) – (See "Hereditary spherocytosis".)

SOCIETY GUIDELINE LINKS

Links to society and government-sponsored guidelines from selected countries and

regions around the world are provided separately. (See "Society guideline links:
Anemia in adults".)

INFORMATION FOR PATIENTS

UpToDate offers two types of patient education materials, "The Basics" and "Beyond
the Basics." The Basics patient education pieces are written in plain language, at the
5th to 6th grade reading level, and they answer the four or five key questions a
patient might have about a given condition. These articles are best for patients who
want a general overview and who prefer short, easy-to-read materials. Beyond the
Basics patient education pieces are longer, more sophisticated, and more detailed.
These articles are written at the 10th to 12th grade reading level and are best for
patients who want in-depth information and are comfortable with some medical
jargon.

Here are the patient education articles that are relevant to this topic. We encourage
you to print or e-mail these topics to your patients. (You can also locate patient
education articles on a variety of subjects by searching on "patient info" and the
keyword(s) of interest.)
 ● Basics topic (see "Patient education: Autoimmune hemolytic anemia (The
Basics)")

SUMMARY AND RECOMMENDATIONS

● Types of hemolysis – Hemolytic anemias can be classified according to whether
the abnormality is intrinsic or extrinsic to the RBC (intracorpuscular versus
extracorpuscular defects), whether the condition is inherited or acquired,
whether the hemolysis is acute or chronic, whether the mechanism involves
antibody-mediated destruction (immune versus non-immune mechanism), and
whether the hemolysis occurs in the vasculature or in the reticuloendothelial
macrophages in the liver and spleen (intravascular versus extravascular
hemolysis). (See 'Conceptual framework' above.)
● Causes – Causes of hemolytic anemia are listed in the table ( table 1) and
discussed above. (See 'List of causes' above.)
● Evaluation – The diagnosis of hemolytic anemia is suspected in a patient with
chronic or new onset anemia with reticulocytosis and not due to another
obvious cause. There is no single specific diagnostic test for hemolytic anemia.
Most experts consider the diagnosis to be accepted if there is reticulocytosis not
explained by recent bleeding, nutrient repletion, or administration of
erythropoietin; with high lactate dehydrogenase (LDH) and unconjugated
bilirubin, low haptoglobin, and in some cases other laboratory findings
( table 2). Once the diagnosis is relatively certain, the cause is determined
using information from the history and physical examination and directed
laboratory testing ( algorithm 1). (See 'Overview of the evaluation' above.)
● Immediate interventions – Certain interventions may be required before the
cause of hemolysis is known, including transfusions, plasma exchange,
hydration, and hemodynamic support. Life-saving interventions should not be
delayed while awaiting the results of diagnostic testing. (See 'Immediate
management issues before the cause is identified' above.)
● Further testing – In some of the obvious/classic presentations, it may make
sense to proceed directly to specific diagnostic testing to determine the specific
cause. If the underlying cause is less obvious, we use the direct antiglobulin
(Coombs) test (DAT) to distinguish between immune and non-immune
 hemolysis, and for those with a negative DAT, we perform directed laboratory
testing based on the patient and family history, physical examination; pace and
severity of hemolysis; and RBC morphology ( algorithm 1). Involvement of
specialists with expertise in hemolytic anemias, laboratory diagnosis, or genetic
testing may be helpful. (See 'Post-diagnostic testing to determine the cause'
above.)
● Atypical presentations – It is possible to have hemolysis without
reticulocytosis, hemolysis without anemia, and reticulocytosis without
hemolysis. (See 'Atypical presentations' above.)
● Thrombosis – There is a well-known association between hemolytic anemia and
thrombosis, which may be seen with either intravascular or extravascular
hemolysis. (See 'Thrombotic complications' above.)

ACKNOWLEDGMENTS

UpToDate gratefully acknowledges Stanley L Schrier, MD (deceased), who

contributed as Section Editor on earlier versions of this topic and was a founding
Editor-in-Chief for UpToDate in Hematology.

The UpToDate editorial staff also acknowledges the extensive contributions of

William C Mentzer, MD, to earlier versions of this and many other topic reviews.
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Topic 7076 Version 70.0

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