wk 3- ab screening Flashcards
serological (hemagglutination-based) tests in immunohematology
advantages
− cost-effective
− highly sensitive
disadvantages
− has many interferences (hemolysis)
− dependent on the ability of immunoglobulin to form complexes or agglutination
- hemolysis is still a reaction (Ab+)
− masked low-incidence Ag
checking for genotype of
patient and donor for
matching (appearance,
genetic code)
molecular method
check for compatibility of donor and patient
blood, presence of agglutination
DIRECT AGGLUTINATION
type of test wherein agglutination means presence of specific Ag
ABO and RH TYPING.
test where agglutination means the donor is not
compatible with patient serum
→ SEROLOGIC CROSSMATCHING.
direct agglutination is used for
→ ABO and RH TYPING. agglutination
means presence of specific Ag
→ detection of IgM
→ SEROLOGIC CROSSMATCHING.
agglutination means the donor is not
compatible with patient serum
. modification of traditional
direct agglutination
GEL TECHNOLOGY
gel technology can be modified using
ahg or antisera
uses
microtiter plates or plates; advantage is 96 tests
can be done in one run
SOLID PHASE RED CELL ADHERENCE
detect the production of
immunoglobulins against patient or donors RBC
COOMBS TEST.
− PRINCIPLE: hemagglutination inhibition
neutralization
− sulfhydryl reagents
→ 2-aminothylisothiouronium bromide
→ 2-mercaptoethanol
→ Dithiothreitol
removal of Ab from RBC to recover
Ab test
ELUTION
acid elution −
change the pH, acidify the pH in order of Ab to elute
o adds glycine-HCl/EDTA
o pH 7.4 – 3 (where Ab reacts)
o buffer solution to neutralize
the pH
heat elution
o submerge the solution to 56Oc
(elute)
o LISS: 37Oc
ui freeze-thaw
elutio
o submerge the samples in -18Oc
then thawing
o centrifuge then collect the
elute
indicates immune-mediated red cell destruction in
the body
DIRECT ANTIGLOBULIN TEST
conditions that results to in vivo sensitization of
RBC (clinically significant)
o HEMOLYTIC DISEASE OF FETUS AND
NEWBORN (HDFN). baby’s blood can
cause sensitization to mother during the
labor
o HEMOLYTIC TRANSFUSION REACTION
(HTR). incompatible transfused blood
o AUTOIMMUNE AND DRUG INDICED
HEMOLYTIC ANEMIA (AIHA). drugs
(rifampin, chloramphenicol), decrease
production of Ab
− detect in vitro sensitization of red cells
INDIRECT ANTIGLOBULIN TEST
o in vivo sensitization of
RBC
dat
in ahg what is the ratio of serum to cells:
1 drop of cell/reagent cell: 2 drops Ab/serum
temp, incubation time, washing and centrifugation in ahg
− temp.: ideal is 37c
− incubation time:
→ LISS: 10-15 mins. to react however, 30-60
mins is common
→ if no reaction enhancement medium,
incubate to 120 mins.
− washing of RBC: 3 times
→ saline washing: should be prepared daily as it can become acidic (acidic environment causes elution)
− addition of AHG
− centrifugation: 2,900 rpm, 3,400 rpm (blood bank)
→ too high causes false+
unexpected Ab
→ immune alloantibodies (primary
importance)
→ naturally occurring alloantibodies
→ passively acquired alloantibodies
→ autoantibodies
test donor/patient serum (unknown) with reagent
RBCs (known)
ANTIBODY SCREENS
reagent vs panel cells
− REAGENT CELLS. Ab detection, set of 2 or 3 vials
− PANEL CELLS. Ab identification, at least 11 vials/set
