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Did You Know? Revisiting Visual Hemolysis in Laboratory Investigation of Transfusion Reaction

A Blog from Eric Ching:

Standard Laboratory Investigation of a Transfusion Reaction entails (AABB Technical Manual):

“ 1. Clerical check of the component bag,label, paperwork, and patient sample.
  2. Repeat ABO testing on the posttransfusion sample.
   3. Visual check of pre- and posttransfusion samples to look for evidence of hemolysis
       (hemolysis may not be visible if <50 mg/dL of hemoglobin is present).
   4. Direct antiglobulin test (DAT) on a posttransfusion sample.
   5. Findings reported to the blood bank supervisor or medical director, who may
       request additional studies or tests.“

Visual hemolysis of the supernatant plasma of the post transfusion sample is the most sensitive and the easiest “test” for  requests of all transfusion reactions to rule out hemolytic transfusion reactions. 

An excellent visual simulation chart of hemolysis can be found in Transfusion 2003;43:297, “a pink hue is  with as little as 2.5 ml of incompatible, lysed RBCs”.

One must compare the appearance of the pre transfusion sample appears to have normal colouration. 

Let’s do a little math: 
Normal plasma HB = 20-30 mg/L (Mollison 5thed., P.566)
Minimum plasma HB to produce visual hemolysis = 200mg/L
 Personal observation for a slight red tinge (using water and EDTA whole blood of known hemoglobin concentration) 250 mg/L 
Difference     250-30 mg/L = 220mg/L
Plasma volume of a 70 Kg patient = 2800mL at 40mL/kg
Amount of HB released from RBC in intravascular hemolysis:  220 mg/L X 2.8L = 616mg
 
Given a normal donor’s MCHC  is 33% or 330 g/L
Assuming the Hct of packed red cells RBC =.75 l/l
The hemoglobin concentration of RBC = 330 g/L x 0.75
                                                                       = 247.5 g/L
Volume of incompatible RCC infused to produce visual hemolysis:
0.616g / 247.6 g/L = 0.00249L or 2.5 mL !
 
Let’s not get too hung up on the numbers as it is essential for us to recognize that it take only a few mL of hemolyzed RBC to give a visual positive test!

However, one must consider other non immune, non pathological causes of hemolysis as well:
  1. Difficult venipuncture
  2. Small bore size of needle 
  3. Accidental over-heat (>47oC) or partially frozen RBC (<-3oC)
  4. Infusion under pressure
  5. ? osmotic damage to red cells
  6. ? infected RBC
Therefore, we must evaluate pre and post transfusion samples along with clinical and laboratory findings before reporting the result. 
 
 Your comments are encouraged!! 

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