ABTIBODY SCREENING

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ABTIBODY SCREENING

• Dr. Mohammed H Saiemaldahr
• BLOOD BANK
• MED TECH DEP
• _at_KAAU

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Antibody Screening

• Antibody Detection and Identification
• Purpose
• The purpose of the antibody screen is to detect
  red blood cell antibodies other than anti-A or
  anti-B.
• These antibodies are called unexpected because
  only 0.3 to 2 of the general population have
  positive antibody screen.
• Once an unexpected antibody is detected, antibody
  identification studies are performed to determine
  the antibodies specificity and clinical
  significance.

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Antibody Screening

• All red blood cell antibodies are significance if
  they cause shortened survival of antigen positive
  red blood cells. For example, anti-D is a
  clinically significant antibody, because it will
  bind to D-positive red blood cells, resulting in
  immune destruction or hemolysis.
• Proper detection and identification of red blood
  cell antibodies is important for the selection of
  appropriate blood for transfusion and in the
  investigation of hemolytic disease of the new
  born, and immune hemolytic anemia.

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Antibody Screening

• Antibody screening test involve testing patients
  serum against two or three reagent red blood cell
  samples called screening cells
• Screening cells are commercially prepared group O
  cell suspensions obtained from individual donors
  that are phenotype for the most commonly
  encountered and clinically important red blood
  cell antigens.

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Antibody Screening

• Group O cells are used so that naturally
  occurring anti-A or anti-B will not interfere
  with detection of unexpected antibodies.
• The cells are selected so that the following
  antigens are present on at least one of the cell
  sample
• D, C, E, c, e, M N, S, s, P, Lea, Leb, K, k,
  Fya, Fyb, and Jkb.

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Antibody Screening

• An anti-gram listing the antigen makeup of each
  cell provided with each lot of screening cells
  issued from a manufacture. It is important that
  the lot number on the screening cells matches the
  lot number printed-On the anti-gram because
  antigen make up will vary with each lot.
• The ideal screening cells have red cells with
  homozygous expression of as many antigens as
  possible.

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Antibody Screening

• There is no requirement that screening cells
  contain red cells with homozygous expression of
  antigens, however, the most workers prefer that
  such red cells are included in screening cells
  sets because many antibodies, especially JK and M
  antibodies, show dosage effect and give stronger
  reactions when tested against cells with
  homozygous expression of their corresponding
  antigen.

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Antibody Screening

• As a result of dosage, weakly reacting antibody
  may not be detected if serum samples are not
  tested against red cells with homozygous
  expression of the their corresponding antigen.

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Antibody Screening

• Screening Cells.
• The screening cells are available in three form
• 1- a single vial of no more than two donors
  pooled together in one vial.
• 2- two vials each with a different donor.
• 3- 3 vials representing three different donors.
• Two or three cells screening sets are required
  for detection of antibodies in pre-transfusion
  testing.

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Antibody Screening

• Detection of very low levels of antibody in a
  recipients serum is important because
  transfusion of antigen-positive red cells may
  result in a secondary immune response with rapid
  production of antibody and subsequent destruction
  of transfused red blood cells.

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Antibody Screening

• Red blood cells should be re-suspended by gentle
  shaking or tilting the tube until the cells no
  longer adhere to the sides. Agglutination is
  graded once the red blood cells are re-suspended.
  
• Agglutination reactions are routinely graded as
  negative (no agglutination).
• Weakly positive, and 1 through 4. The degree of
  the positive reaction generally indicates the
  amount of Ab present not its significance.

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Antibody Screening

• Auto-logous Control.
• Autologous control is considered as part of the
  Ab screening, it can be performed in parallel
  with the Ab screen and involves testing the
  patients serum against the patients red blood
  cells.
• A positive auto-logous control is an abnormal
  finding and usually means that patient has a
  positive direct antiglobulin test (DAT).

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Antibody Screening

• Grading Reactions.
• Aggregation or hemolysis of test red blood cells
  is the visible end point of an Ab-Ag interaction.
  
• Test results should be read immediately after
  centrifugation as delays in reading may cause
  elution of antibody and false- negative test
  results
• The first step in reading hem-agglutination
  reactions is inspection of the supernatant for
  signs of hemolysis (red or pink coloration).

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Antibody Screening

• Interpretation.
• Evaluation of the antibody screening and
  auto-logous control results can provide clues and
  give direction for the identification and
  resolution of the Ab or Abs. The investigator
  should consider the following questions

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Interpretation

• 1- In what phase (s) did the reaction (s) occur?
• Low temperature Abs of the IgM class will react
  best at low temperatures and are capable of
  causing agglutination of saline-suspended red
  blood cells (immediate spin reading (IS). Of the
  commonly encountered Abs are, anti-N, -I. and -P
  (IgM)
• Abs of the IgG class will react best at the AHG
  Phase (37C). Of the commonly encountered Abs
  are anti-Rh, kell, Kidd, and Duffy (IgG).
• Where as Lewis, M, Abs may be IgG, IgM, or a
  mixture of both.

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Interpretation

• 2- Is the auto-logous control negative or
  positive?
• A positive Ab screen and a negative auto-logous
  control indicate an allo-antibodv has been
  detected.
• A positive auto-logus control may indicate the
  presence of auto-antibodies, antibody to
  medication or it may idiopathic
• If the patient has been recently transfused, the
  positive auto-logous control may be due to
  alloantibody coating circulating donor red blood
  cells.

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Interpretation

• 3- Did more than one screening cell react and if
  so, did they react at the same strength and
  phases?
• If the patient has multiple Abs, when the Abs
  corresponding Ag is found on more than one
  screening cell, or when the patients serum
  contains an autoantibody, more than one screening
  cell will be positive.
• A single Ab specificity should be suspected when
  all cells react at the same phase and strength.
• Multiple Abs are most likely when cells react at
  different phases and strengths and
  auto-antibodies are suspected when the
  auto-logous control is positive.

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Interpretation

• 4-Are the cells truly agglutinated or rouleaux
  present?
• Serum from patients with multiple myeloma or who
  have received high molecular weight plasma
  expanders (dextran) may cause non-specific
  aggregation of red blood cells known as rouleaux.
• Rouleaux is not a significant finding in Ab
  screening tests but it is easily confused with Ab
  mediated agglutination. To differentiate between
  rouleaux and agglutination

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Interpretation

• 1- Rouleaux cells have a stacked coin
  appearance when viewed microscopically.
• 2- It will not interfere with the AHG phase of
  testing because the patients serum is washed
  away prior to the addition of the AHG reagent.
• 3- It is observed in Ab tests containing the
  patients serum including the auto-logous control
  and the reverse ABO typing.
• 4- Rouleaux is dispersed by the addition of 1 to
  3 drops of saline to the test tube.

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Reagent Red Blood Cell Screening Cell Sectional
Listing of Antigens Present
CC AHG 37 IS P1 N M s S Leb Lea Jkb Jka Fyb Fya k K e c E C D No cell
2 0 0 0 0 0 0 0 I
2 0 0 0 0 0 0 0 0 0 0 0 II