Trend

in Laboratory Diagnostic
for Allergy

DEWI WULANDARI
Dept. of Clinical Pathology
Faculty of Medicine, University of Indonesia
Dr. Cipto Mangunkusumo Hospital
Jakarta
 Introduction
Immune system :
• Protection against
any potentially
harmful agents.
• No significant injury
on host cells or
tissues
 Hypersensitivity Reactions
• Against normally
harmless agents.
• Results in host cells
or tissue injury 
diseases
• Manifestations :
Depend on mediators
involved
Tissues affected
 Classification of Hypersensitivity
Reactions
(Coombs & Gel -1963)

Type I Type II Type III Type IV

Immediate Antibody mediated Immune complex T-cell mediated

hypersensitivity diseases mediated disease hypersensitivity

IgE mediated IgG & IgM against cell IgG & IgM against Delayed-type
surface or circulating antigen hypersensitivity
Mast cells, basophils,
extracellular matrix
eosinophils Deposition of
immune complex

Abbas AK, et al. Basic Immunology: Functions and disorders of the immune system. 4th ed.
 Allergens
• Antigens that elicit
allergic reactions
• Normally harmless
• Thousands of allergens
have been identified
• Sequenced allergen
 recombinant protein
 Immunoglobulin E
• Prausnitz & Küstner (1921) : discovered
IgE involvement in allergic reactions.

• IgE mediated inflammatory :

– Part of immune response to parasites
– Immediate-type hypersensitivity
• Highest concentration of IgE-producing
plasma cell : lymphoid tissue associated
to GI & respiratory tract.
 Immunoglobulin E
• The lowest of Ig isotypes: 0.002% of total serum
immunoglobulin
• IgE has the highest affinity for antigens and for its
receptors.
• The tremendous biologic amplification linked to IgE
and its receptors is shown by the fact that even low
(ng/mL) concentrations of cell surface IgE can cause
life-threatening reactions within minutes of allergen
exposure in highly sensitive individuals.

• Short half-life : 1-5 days.

Smith PH, et al. Middleton’s Allergy: principles and practice 8th ed.
 TOTAL IgE LEVEL – Age-related

Middleton's Allergy: Principles and Practice 8th Edition

Copyright © 2014
Middleton's Allergy: Principles and Practice 8th Edition
Copyright © 2014
Diagnosis of Allergic Diseases
• Clinical history
• Physical examination
• Additional Examinations
– Total IgE antibody
measurement
– Allergen-specific IgE
antibody tests
– Allergen provocation test
 Total IgE antibody Measurement
• Purpose :
– to screen out non-IgE related clinical conditions
– (Anti IgE antibody treatment monitoring)
• Interpretation : cut-off 100 IU/ml
• Test principle

© 2012 Thermo Fisher Scientific Inc.

Allergen-specific IgE antibody Tests

• To confirm allergen
specificity
• Useful for :
– Allergen avoidance
– Allergen desensitization
• Methods :
– In vivo (skin prick tests)
– In vitro (immunoassays)
 Allergen-specific IgE antibody Tests
Skin prick tests vs Immunoassays

Immunoassays Skin Prick Tests

Lack of risk for allergic reaction Greater sensitivity
Results unaffected by drugs Results immediately available
Results unaffected by conditions of Less time and reagent expenses
skin
Greater patient convenience
Special expertise not required
Easier-to-document QC
Well standardized and reproducible
Costly instrument and reagent Need antihistamine cessation
Inapplicable in patient with skin
problem
Risky in uncooperative patients
Smith PH, et al. Middleton’s Allergy: principles and practice 8th ed..
 Allergen-specific IgE antibody
Immunoassay
Single-allergen assays vs Multiplex-allergen assays

© 2012 Thermo Fisher Scientific Inc.

 EXTRACT vs MOLECULAR ALLERGENS

The most
allergenic protein

Molecular
component used
in ISAC

Other specific IgE directed against

lower risk peanut proteins such as Ara
h 8 and Ara h 5. Proteins may cross
react with other sources

extract-based IgE blood tests measure the “sum” of molecular allergology makes it possible to investigate important individual proteins
sensitization to all protein components in whole within a peanut for specific IgE sensitization.
allergens, e.g. peanut,
EXTRACT ALLERGENS

• Extract allergens includes

all protein component in
whole allergens
• Specific IgE levels do not
always correlate with
clinical symptoms
MOLECULAR COMPONENT ALLERGENS
• Molecular-based allergy diagnostics is used
to map the allergen sensitization at a
molecular level
• increased accuracy in allergy diagnosis and
prognosis
• Has important role in:
– resolving genuine versus cross-reactive
sensitization
– assessing the risk of severe, systemic vs
mild, local reactions
– identifying patients and triggering
allergens for specific immunotherapy
(SIT).
MOLECULAR COMPONENT ALLERGENS
Allergen components are named:
• based the Latin name of the allergen source
• and a number based on the order of discovery
(when registered/ approved by the IUIS
committee)

example of peanut allergen component nomenclature:

Peanut – Arachis hypogaea – Ara h 2
MOLECULAR COMPONENT ALLERGENS
MOLECULAR COMPONENT ALLERGENS

High- versus low-risk molecules from foods

giving rise to anaphylaxis

Canonica et al. World Allergy Organization Journal 2013, 6:17

Single Allergen vs Multiplex Immuno Solid-phase Allergen Chip
(ISAC)
 BASOPHIL ACTIVATION TEST

• In vitro test to measure basophil activation upon

allergen stimulation
• Functional assay to test functional specific IgE
crosslinking
• Can be part of diagnostic tools, monitoring for
allergen immunotherapy
BASOPHIL ACTIVATION TEST

Santos and Lack Clin Transl Allergy (2016) 6:10

General diagnostic work up of IgE mediated diseases
 SUMMARY
• Allergy is currently a public health issue although hugely underestimated and under-
recognized.
• Diagnosis of allergy are based on careful history taking and physical examination,
supported by laboratory tests.
• Total serum IgE level may differentiate between allergy and non-allergy conditions.
• Specific allergy testing are used to identify patient sensitization as a basis for specific
immunotherapy or allergen avoidance.
• Molecular component are currently used for specific IgE testing replacing allergen
extract
• Basophil activation test is a functional IgE test entering clinical application