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CLS 3311 Advanced Clinical Immunohematology

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CLS 3311 Advanced Clinical Immunohematology Antihuman Globulin Follow-Up Lecture False Positive: IAT or DAT Tubes or saline contaminated with excess silicone Dirty ... – PowerPoint PPT presentation

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Title: CLS 3311 Advanced Clinical Immunohematology


1
CLS 3311Advanced Clinical Immunohematology
  • Antihuman Globulin Follow-Up Lecture

2
False Positive IAT or DAT
  • Tubes or saline contaminated with excess silicone
  • Dirty glassware
  • Overcentrifugation
  • Over reading
  • May cause non specific aggregation
  • May cause non specific aggregation
  • RBCs tightly packed results in difficulty
    dispersing can mistake for agglutination
  • Just looking too close

3
False Positive IAT or DAT
  • Polyagglutinable RBCs
  • Inherited exposure of T, Tn, Tk antigens, also
    known as crypt antigens that agglutinate with
    virtually all patient serum.
  • Using contaminated anti-serum
  • Patient has infection with enzyme secreting
    bacteria, the enzymes strip sialic acid from cell
    surface exposing crypt antigens.
  • Chapter 22, Harmening

4
False PositiveDATs
  • Complement component C4 may bind to red cell
    membrane of clotted specimens at 4?C and Room
    temperature
  • May even occur in CPDA-1 (donor anticoagulant)
    red blood cells
  • can be avoided by using anticoagulant that
    chelates Ca such as EDTA

5
False PositiveDATs
  • Blood collected from IV lines used to infuse 5 or
    10 dextrose in H2O or volume expanders
  • dextrose causes cells to aggregate
  • Septicemia or bacterially contaminated sample
  • bacterial enzymes remove sialic acid from RBCs
    causing exposure of T antigens

6
False Negative IATs DATs
  • Associated with Washing
  • Cells NOT washed adequately
  • Inadequate resuspension of RBCs between washes
  • Contamination of cells during washing
  • Interruption, leaving cells in saline for
    prolonged period - Antibody elutes off of red
    blood cell

7
False Negative IATs DATs
  • Associated with AHG Reagent
  • Deterioration of AHG in storage
  • Failure to add AHG to test
  • Delay in adding AHG to washed cells

8
False Negative DAT
  • Improper centrifugation
  • Undercentrifugation agglutination may be missed
  • Overcentrifugation fragile agglutination may be
    broken up trying to pry cells off tube
  • Too many RBCs
  • Ag excess suspension too heavy - difficult to
    read
  • Too few RBCS
  • Too many cells lost during Washing step
  • Patient RBC suspension too weak

9
False Negative DAT
  • Monoclonal gammopathies
  • IgG concentration so high even 4 washes dont
    completely remove free IgG - neutralize anti-IgG
  • Less than 200 IgGs attached per RBC
  • IgG concentration so low theres not enough to
    attach onto red cell membrane to detect their
    presence

10
False Negative IAT
  • RBCS and/or serum stored improperly
  • Can cause loss of RBC antigens
  • Can cause reduction of antibody activity
  • Presence of Antibody that does not react with
    monoclonal anti-IgG
  • Can only be detected with anti-C or may be IgA

11
False Negative IAT
  • Improper incubation
  • Temperature not at 37 ?C (/ 1oC)
  • Incubation time not adequate for agglutination to
    form
  • Incubation time so long, Antibodies begin to
    dissociate
  • Patient serum not added
  • Poor reading technique
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