Antibody Detection and Identification

0.0(0)
studied byStudied by 2 people
learnLearn
examPractice Test
spaced repetitionSpaced Repetition
heart puzzleMatch
flashcardsFlashcards
Card Sorting

1/38

encourage image

There's no tags or description

Looks like no tags are added yet.

Study Analytics
Name
Mastery
Learn
Test
Matching
Spaced

No study sessions yet.

39 Terms

1
New cards

Antibody Screen

  • Test used to detect Abs

  • Used pts plasma or serum against reagent RBCs to detect unexpected Abs

  • Unexpected Abs are a result of RBC stimulation

    • Transfusion

    • HDFN

2
New cards

When are Antibody Screens used?

  • Pts needing a transfusion

  • Pregnant women

  • Pts who have had transfusion rxns

  • Blood and plasma donors

3
New cards

Which unexpected Ab is clinically significant?

  • Clinically significant (IgG)

  • Not clinically significant (immunoglobulin M [IgM]

4
New cards

Clinically Significant Antibodies

  • Usually IgG

  • React best at 37° C and during the antihuman globulin (AHG) phase (indirect antiglobulin test [IAT])

  • are associated with hemolytic transfusion reactions

    (HTRs) and HDFN

5
New cards

Performing an Antibody Screen

  • pt’s plasma or serum is incubated with screening cells

  • After incubation, an IAT is performed using AHG reagent

  • Want to detect any IgG antibodies

6
New cards

Screening Cells

  • are single or pooled donor group O cells

    • however, single-donor vials offer increased sensitivity

  • Group O cells are used so that anti-A and anti-B antibodies will not react

  • Screening cells come in sets of 2 or 3 vials (donor) each

    • Each vial has been phenotyped for each antigen

7
New cards

Antigram

A reaction to one or more cells indicates the presence of an atypical antibody,

8
New cards

Autocontrol

Tests a patient’s serum with his or her own RBCs

  • incubated with the Ab screen (or Ab panel)

  • if AC w/ a screen and it is (+), can run a DAT (patient cells plus AHG) to detect in vivo coating

    • help determine if Abs are directed against the pt’s cells or transfused cells (allo- or autoantibody)

9
New cards

Potentiators

Used in Ab detection and identification to enhance an Ag–Ab rxn

  • Saline (may only enhance if incubated for a long time)

  • Low-ionic-strength solution (LISS): common

  • Bovine serum albumin (BSA)

  • Polyethylene glycol (PEG)

  • Proteolytic enzymes (can destroy some antigens)

10
New cards

Comparison of Potentiators and Methods

11
New cards

Patient History Significance

GET THE HISTORY!

  • Mixed RBC pop. from a prev. transfusion can remain for up to 3 months

  • Pt may have come from another hospital

  • Some diseases are associated with Abs

  • Some Abs occur at a higher frequency in some races

  • Learn about the diagnosis, age, and race of the patient

12
New cards

Antibody ID: Initial Panel

an extended version of an antibody screen (10 to 20 cells)

  • Group O cells

  • Each panel cell is antigen typed

  • its antigram lists the phenotypes of each panel cell

    • used to record and interpret results

13
New cards

Interpretation Guidelines

  • Autocontrol: A (+) autocontrol suggests autoantibodies or alloantibodies to transfused cells, while (-) result = alloantibodies.

  • Phases: IgM reacts at room temp/IS phase, IgG reacts at 37°C/AHG phase.

  • Reaction Strength: Varying strengths suggest multiple Abs or dosage effects.

  • Ruling Out Antibodies: Use fully (-) panel cells to rule out Abs, avoiding heterozygous exclusions.

  • Matching the Pattern: A single antibody shows a clear pattern; multiple Abs show mixed patterns.

  • Rule of Three: Confirm AB ID with 3 antigen-positive and 3 antigen-negative cell reactions.

14
New cards

Autocontrol (AC) interpretation

  • Positive autocontrol or DAT:

    • Autoantibody or alloantibody present to recently transfused cells

  • Negative autocontrol: indicates alloantibody

  • Positive autocontrol + negative DAT:

    • Indicates false positive test

15
New cards

Phases Interpretation

  • IgM Abs react at room temp. or during immediate-spin (IS) crossmatching

    • anti-Lea, anti-Leb, anti-M, anti-N, anti-I, anti-P1 Abs

  • IgG antibodies usually react at 37° C or with AHG

  • Rxn at diff phases may indicate IgM AND IgG

16
New cards

Reaction strength Interpretation

  • Rxns w/ varying strengths can indicate multiple Abs

  • Strength also affected by dosage

17
New cards

Ruling Out Antibodies Interpretation

  • Use panel cells (-) in all phases to rule out Abs.

  • Start with the first (-) panel cell and cross out present Ags.

  • Do not cross out heterozygous panel cells; Abs may be too weak to react.

18
New cards

Matching the Pattern Interpretation

  • pattern of rxns matches one of the Ag columns when a single Ab is present

  • Multiple Abs show varying patterns

19
New cards

Rule of Three Interpretation

  • To ensure valid results, 3 Ag (+) cells must react and 3 Ag (-) cells must not react with the pt’s serum or plasma

  • If this does not occur, additional panel cells (selected cells) are used

20
New cards

Example of Ruling Out and Matching the Pattern

21
New cards

Phenotyping the patient

  • confirm the presence of Abs is to determine the pt’s phenotype for Ags

  • Individuals do not make alloantibodies toward Ags on their own RBCs

  • If the patient has recently been transfused, RBC separation techniques should be used before phenotyping is done

22
New cards

Multiple Antibodies

Special techniques can be used to help identify multiple Abs

  • Selected cells

  • Proteolytic enzymes

  • Other chemicals (e.g., dithiothreitol)

  • Dithiothreitol = denature Kell antigen

23
New cards

Proteolytic Enzymes

used to eliminate or enhance Ab activity

  • Duffy and MNS Ags are destroyed

    • Decrease

  • Rh, Kidd, and Lewis antigens are enhanced

    • Increase

  • One-stage test: Enzymes, RBCs, and serum are simultaneously incubated

  • Two-stage test: Panel cells are pretreated with enzymes, washed, and then used as usual

24
New cards

Proteolytic Enzymes Examples

include papain, bromelain, and ficin, used to alter antigen expression on red blood cells.

<p>include papain, bromelain, and ficin, used to alter antigen expression on red blood cells. </p>
25
New cards

Antibodies to High-Frequency Antigens

  • High-frequency Ags occur in the pop. at a frequency of 98% or higher

  • Suspect an alloantibody to a high-frequency Ag if most panel cells are positive

  • Testing to confirm include:

    • Enzymes to enhance or destroy Ags

    • Dithotheritol to destroy Kell Ags

<ul><li><p>High-frequency Ags occur in the pop. at a frequency of 98% or higher</p></li><li><p>Suspect an alloantibody to a high-frequency Ag if most panel cells are positive</p></li><li><p>Testing to confirm include:</p><ul><li><p>Enzymes to enhance or destroy Ags</p></li><li><p>Dithotheritol to destroy Kell Ags</p></li></ul></li></ul><p></p>
26
New cards

Clues for Identifying High Frequency Antibodies

27
New cards

High-Titer, Low-Avidity Antibodies

Antibodies to high frequency antigens that react weakly

  • React at AHG phase

  • Not implicated in transfusion reactions of HDFN

<p>Antibodies to high frequency antigens that react weakly </p><ul><li><p>React at AHG phase</p></li><li><p>Not implicated in transfusion reactions of HDFN</p></li></ul><p></p>
28
New cards

Anitbodies to Low-Frequency Antigens

  • may occur alone or suspect when screen is negative and crossmatching is positive

  • one one reactive cell suggest this

<ul><li><p>may occur alone or suspect when screen is negative and crossmatching is positive </p></li><li><p>one one reactive cell suggest this</p></li></ul><p></p>
29
New cards

Enhancing Weak IgG Antibodies

30
New cards

Cold Alloantibodies

  • IgM antibodies that react during IS crossmatching (sometimes 37 C)

  • Not clinically significant

  • Anti-P, anti M, and anti-N antibodies have variable rxns

    • to enhance rxns: incubation < 37 C

    • to avoid rxns: neutralize to allow for detection of clinically sig, Abs

<ul><li><p>IgM antibodies that react during IS crossmatching (sometimes 37 C) </p></li><li><p>Not clinically significant</p></li><li><p>Anti-P, anti M, and anti-N antibodies have variable rxns</p><ul><li><p>to enhance rxns: incubation &lt; 37 C</p></li><li><p>to avoid rxns: neutralize to allow for detection of clinically sig, Abs</p></li></ul></li></ul><p></p>
31
New cards

Autoantibodies

  • react w/ all reagents & self & donor RBCs, regardless of present Abs

  • AC & DAT (+)

  • must identify underlying AlloAb when autoAbs present

  • Adsorption used for removal of AutoAbs

32
New cards

Cold Autoantibodies

  • React during IS crossmatching

  • Positive autocontrol and DAT (to C3)

  • May occur with:

    • Mild anemia

    • Mycoplasma pneumoniae infection

    • Infectious mononucleosis

  • Most are anti-I, anti-H, and anti-IH

    • "Cold panels" can aid identification

<ul><li><p>React during IS crossmatching</p></li><li><p>Positive autocontrol and DAT (to C3)</p></li><li><p>May occur with:</p><ul><li><p>Mild anemia</p></li><li><p>Mycoplasma pneumoniae infection</p></li><li><p>Infectious mononucleosis</p></li></ul></li><li><p>Most are anti-I, anti-H, and anti-IH</p><ul><li><p>"Cold panels" can aid identification</p></li></ul></li></ul><p></p>
33
New cards

Avoiding Cold Autoantibody Reactions

  • Use a monospecific IgG AHG reagent rather than a polyspecific reagent

  • Skip IS crossmatching and testing at 37° C

  • Use 22% bovine serum albumin instead of LISS

  • Prewarming all tubes to 37° C will avoid reactivity

    • Washing with warm saline is not advised when prewarming the tubes

  • Use adsorption techniques if all else fails

34
New cards

Adsorption techniques

35
New cards

Warm Autoantibodies

  • More common than cold autoantibodies

  • May result from autoimmune hemolytic anemia, disease, or drug.

  • Most panel cells and the AC are (+)

  • 22% albumin decreases reactivity

  • Determine if underlying alloantibodies exist

36
New cards

Specificity

  • Warm autoantibodies often target Rh system, esp. e antigen.

    • Patient has anti-e antibody and e antigen

    • Positive DAT.

    • e-negative blood may improve RBC survival in chronic hemolysis.

  • Medications can induce warm autoantibodies.

37
New cards

Drug Mechanisms Associated w/ Positive DAT

38
New cards

What is Elution? What are its methods?

  • Used to identify antibodies when the DAT is positive.

  • IgG must be detached by using the elution technique

  • The recovered antibody is called the eluate

  • Eluate is used in an antibody panel to identify antibody

  • Elution is performed in suspected cases of HDFN; may not be reactive in all cases of warm autoantibodies

<ul><li><p>Used to identify antibodies when the DAT is positive.</p></li><li><p>IgG must be detached by using the elution technique</p></li><li><p>The recovered antibody is called the <mark data-color="yellow" style="background-color: yellow; color: inherit">eluate</mark></p></li><li><p>Eluate is used in an antibody panel to identify antibody</p></li><li><p>Elution is performed in suspected cases of HDFN; may not be reactive in all cases of warm autoantibodies</p></li></ul><p></p>
39
New cards

Adsorption

  • Remove warm autoantibodies to test for underlying alloantibodies

  • Cells pretreated with dithiothreitol or ZZAP to remove in vivo attached antibodies