174) Laboratory test for iron stores & availability
Iron; needed for life, but potentially very toxic (free radicals)
Tests of iron metabolism
Serum iron ( SI)
- F: 600-1400 mg/L, 11-25mmol/L
- M: 750-1500 mg/L, 13-27mmol/L
- Low in Fe deficiency and chronic disease
- High in hemolytic syndromes and iron overload
Total iron binding capacity (TIBC)
- amount of iron needed to bind to all the transferrin
- 2500 – 4500 mg/L , 45-82 mmol/L
- High in Fe deficiency
- Low in chronic disease
Serum ferritin
- Fe storage glycoprotein
- Can store up to 2000 Fe
- (30-300 ng/mL)
- Serum level is very low, but closely correlates with level in cells
- Closely correlates with total body Fe stores
- <12 ng/mL Fe deficiency
- Elevated in Fe overload, liver injury, tumors (Acute phase protein)
- Tests for iron metabolism
Additional;
Serum transferin receptor
Increase in increased erythropoiesis and early Fe deficiency
RBC ferritin
storage status over the previous 3 month (Fe deficiency/overload)
unaffected by liver function or acute illness
Free RBC porphyrin
increased when heme synthesis altered
Iron Deficiency Anemia
Prelatent; (Decreasing iron stores of organism)
- Decrease in serum ferritin – most sensitive parameter
- decrease of iron in BM – (iron is in BM cells in form of ferritin)
- increase TIBC (body has tendency to increase the absorption of iron and iron transporting
capacity) this leads to decrease in Tf saturation even when serum iron is normal
Latent; (Decreases serum iron available for erythropoiesis)
- decrease serum iron
- further decrease Tf saturation
- increase sTfR
Manifest anemia
- parameters of anemia (low Hb, Hct, Erythrocyte count)
- anemia of iron deficiency is hypochromic and microcytic
Anemia of chronic
diseases
Anemia due to lack of
iron
Myelodysplastic
syndrome
Iron is locked into
macrophages to be out
of reach of bacteria
Lack of stores, see
prelatent, latent and
manifest anemia
Aplastic disorder of
BM, enough iron in
organism
Serum iron
Transferrin/TIBC
ferritin
References:
175) Blood tests preceding blood transfusion. Laboratory
indicators of hemolysis.
1. Blood tests preceding blood transfusion
Indications of blood transfusions:
-Massive blood loss due to trauma or surgery.
-Severe anemia or trombocytopenia
-Hemophilia, sickle-cell disease – often frequent transfusions.
-Need for blood based products – e.g. factor concentrates in Liver cirhosis or Immunoglobulins in some
immunodeficiency states.
Two types of transfusion:
Homologous (allogenic) transfusion - using blood or blood products of others.
Autologous transfusion - using the patient's own stored blood (planned surgery, less risk of infection).
Blood products currently used in transfusions:
Whole blood
Red blood cells
Plasma or Fresh Frozen Plasma (FFP)
Platelets /these 3 can be obtained from the whole blood by centrifugation
Albumin protein
Cryoprecipitate (prepared from plasma by cryoprecipitation (“freezing out”
some proteins; it
contains f. VIII, fibrinogen, vWfactor, f. XIII. It can be used e.g. in Hemophilia, vonWillebrandt disease,
DIC and hypofibrinogenemia)
Clotting factor concentrates,
Fibrinogen concentrate
Immunoglobulins (antibodies)
Blood groups:
The most important blood groups are of AB0 system (incompatibility can cause hemolysis after
transfusion, there antibodies are IgM and thus are big enough to cause agglutination) and Rh system
(incompatibility between mother and child causes hemolytic disease of the newborn). However there are
more then 30 other groups of antigens present on the surface of RBC. To give some examples: MNS
system (especially anti-S can cause hemolysis) and Kell antigen system (incompatibility can cause AIHA
and hemolytic disease of the newborn). Presence of antibodies against these antigens (other then ABO
and Rh system) in plasma is rare but if they appear, they can cause serious transfusion reactions. These
antibodies are called irregular antibodies.
Testing done in laboratory prior to the transfusion:
1) Donor’s and Patient/recipient’s blood group (AB0 and Rh system)
2) Screening of the recipients serum for the irregular antibodies (this is done by mixing patients
serum with standardized mixtures of erythrocytes that contain all known irregular antigens.
Agglutination means presence of irregular antibodies and calls for further more detailed
identification of the type of the irregular antibody. This should be done every day of transfusion,
as new antibodies can appear as a reaction to previous transfusion!
3) Cross-matching: Red blood cells from the donor unit are tested against the plasma of the
patient in need of the blood transfusion (big cross-matching examination). If the patient’s serum
contains antibodies against the antigens present on the donor red blood cells, agglutination will
occur. Agglutination is considered a positive reaction indicating that the donor unit is
incompatible for that specific patient. If no agglutination occurs the unit is deemed compatible
and is safe to transfuse. In some countries, cross matching examination is not done, when there
are no irregular antibodies present in recipient serum. In Czech Rep. it is always done.
4) Screening for infectious diseases A number of infectious diseases can be transmitted from donor
to recipient. In order to avoid this, screening for potential risk factors and health problems among
donors is done and laboratory testing of donated units for infection is a standard procedure in
developed countries. Diseases that can be transmitted: HIV, Hepatitis A,B and C, Human Tlymphotropic virus, West Nile virus, Treponema Pallidum, Malaria, Chagas disease,
Cytomegalovirus.
Bed side testing
1) Doctor (in Czech lands) or nurse checks the patients name and number on the blood product and
the documentation to avoid confusion.
2) Bed side paper cross-matching examination. Two drops of the patient’s blood are mixed on a
piece of paper with AntiA and AntiB reagents (Antibodies). Agglutination determines the blood
group of the patient (e.g. if aglutination appears with Anti A, blood group is A). Below that on
the same piece of paper, same is done with the donated blood from the bag. The blood groups
have to be same.
3) Biological test – patient is infused blood for 5 minutes. Then, transfusion is paused for 15
minutes. Meanwhile, we watch for nausea, fever etc. If everything is ok, we continue. This is not
done in urgency.
Note: In urgent cases, be can give the blood of group 0 (from so called universal donor) to any
recipient.
Types of transfusion reactions:
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Febrile non-hemolytic transfusion reaction
Bacterial infection
Acute hemolytic reaction
Anaphylactic reaction
Transfusion-associated acute lung injury (TRALI)
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Volume overload
Iron overload – in frequent transfusions
Delayed haemolytic reaction
Transfusion-associated graft vs. host disease (GVHD) – in immunodeficiency
2. Laboratory indicators of hemolysis
Hemolysis – destruction of red blood cells and release of haemoglobin into the surrounding fluid. In
extravascular hemolysis (e.g. in hemolytic anemia), the red blood cells are destroyed by macrophages in
organs like spleen and liver and hemoglobin is not released into blood stream. In intravascular
hemolysis (e.g. after incompatible transfusion), RBC are destroyed inside the blood vessels. Hemoglobin
is released into blood stream, where it binds with haptoglobin. Unbound hemoglobin or heme is filtrated
into urine and can cause iron damage to the cells of proximal tubule.
Haptoglobin - protein in the blood plasma that binds free hemoglobin released from erythrocytes with
high affinity and thereby inhibits its oxidative activity. Its level decreases (it is spent) after intravascular
hemolysis.
Haemopexin – Binds and scavenges potentially toxic heme, preventing its release into urine. Low levels
are indicative of hemolysis.
Schumm test:
The Schumm test (shoom) is a blood test that uses spectroscopy to determine significant levels of methaemAlbumin in the blood. A positive
result could indicate intravascular hemolysis.
A positive test result occurs when the haptoglobin binding capacity of the blood is saturated, leading the hemoglobin to bind to albumin instead.
Further excess in hemoglobin will result in hemoglobinemia and hemoglobinuria. Hemosiderinuria will also result if there is chronic intravascular
hemolysis.
Unconjugated Bilirubin – After breakdown of heme, the unconjugated bilirubin is released into blood
stream by macrophages. It is normally conjugated in liver and released into bile. If there is more
breakdown of heme, there will be more unconjugated bilirubin and jaundice can appear. High levels of
unconjugated bilirubin are indicative of substantial hemolysis.
Other testing:
Coombs test (direct antiglobulin test):
Used to detect IgG antibodies or complement proteins that are bound to the surface of red blood cells;
RBCs are washed (removing the patient's own plasma) and then incubated with antiglobulin (also known
as "Coombs reagent"). If this produces agglutination of RBCs, the direct Coombs test is positive,
indication that antibodies (and/or complement proteins) are bound to the surface of red blood cells.
Important test in the diagnosis of AIHA (autoimmune hemolytic anemia).
Test of osmotic resistance (sugar or sucrose lysis test):
Patient's red blood cells are placed in low ionic strength solution and observed for hemolysis.
Ham’s acid hemolysis:
Specific test to diagnose Paroxysmal nocturnal hemoglobinuria (PNH). Low pH activates complement
system. RBCs in PNH are more susceptible to complement damage.
More sensitive modern methods include flow cytometry for CD55 and CD59 on white and red blood
cells (these are absent or low in PNH)
References: