EBV STEPWISE DIAGNOSIS STEP ONE EBNA-1-IGG-ELISA PKS MEDAC - EBV EBNA-1 MEDAC
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medac
Diagnostika
EBV EBNA-1
EBV Stepwise Diagnosis
Step One
EBNA-1-IgG-ELISA PKS medac
medac
Gesellschaft für klinische Spezialpräparate mbH GE Diagnostika Theaterstrasse 6 D-22880 Wedel
Telefon 04103/8006-0 Fax 04103/8006-359 www.medac.demedac
EBV EBNA-1: IgG Serology from medac
Importance
Epstein-Barr virus (EBV), a member of the Herpesviridae family, consists of a
double-stranded DNA genome, a capsid, matrix and virus envelope.
Following primary infection, EBV typically persists in latent form in the body
for the rest of the individual's life.
Between 90% and 95% of all adults worldwide have been infected with EBV.
In immunocompetent individuals primary EBV infections usually follow an
asymptomatic course in early childhood. Even minor immunosuppression
may cause the virus to be reactivated although this is not usually
accompanied by clinical symptoms.
Antibody assays are used primarily for EBV diagnosis in immunocompetent
individuals. In diagnostic terms, the relevant antigen complexes are EBV-
specific nuclear antigen (EBNA-1), virus capsid antigen (VCA) and, where
appropriate for differential diagnostic enquiries, early antigen (EA). In this
context, the detection of EBNA-1 antibodies should always take
precedence.
The presence of EBNA-1 antibodies invariably testifies to previous EBV
infection. A negative EBNA-1 result is the starting point for differential
diagnosis employing further serological investigations.
Disease
The following diseases are directly or indirectly related to EBV and indicate
associations serological investigation:
- Infectious mononucleosis
- EBV-associated B-cell lymphoma
- Burkitt's lymphoma (endemic in Central Africa and New Guinea)
- Nasopharyngeal carcinoma (increased incidence in Southeast
Asia and China)
- Hodgkin's lymphoma
2medac
EBV EBNA-1: IgG Serology from medac
Antibody
diagnosis The primary goal of EBV antibody diagnosis in immunocompetent
individuals is to differentiate between:
- primary infection
- seronegative status and
- previous infection
Antibody
pattern in
primary Primary EBV infection is characterised by the following typical antibody
response pattern:
infection
Symptoms
Infection
VCA IgG
VCA IgA
EBNA-1 IgG
VCA IgM
EBNA-2 IgG
EA IgG
3medac
EBV EBNA-1: IgG Serology from medac
Efficient
stepwise To encourage efficient serological testing for EBV, the National Reference
diagnosis Laboratory for EBV (Virology Institute, University of Homburg/Saar)
recommends the following diagnostic approach. Further diagnostic steps
are necessary only if EBNA-1-IgG is not detected. This diagnostic approach
is informative as well as efficient in terms of time and cost.
EBNA-1IgG
positive negative
previous infection VCA IgG
positive negative
VCA IgM seronegative
positive negative
Further diagnostic steps,
primary infection
e.g. avidity, blot, PCR
4medac
EBV EBNA-1: IgG Serology from medac
Serological
results Determination of EBV antibodies in immunocompetent individuals generally
permits unequivocal interpretation of serological status (see table).
EBNA-1 IgG VCA IgG VCA IgM Interpretation
- - - seronegative
- + (-) + Primary infection /
Reactivation
+ + - Previous
infection
+ + + ?*
- + - ?*
* EBNA-1 antibodies are not detectable in about 5% of patients with
previous EBV infection.
In the event of a negative result for EBNA-1-IgG, an isolated positive
result for VCA-IgG or general VCA-IgG/-IgM and EBNA-1-IgG positive
status, further differential diagnostic steps are necessary
(determination of heterophile antibodies, avidity, measurement of
p18, PCR).
Advantages
of the Binding assays such as IFT and ELISA are routinely used for the detection of
assay EBV-specific antibodies.
- EBNA-1-IgG-ELISA-PKS medac detects antibodies directed
specifically against the nuclear antigen.
- The assay is quantitative.
- It implements the principle of single-point quantification.
- The assay has been CE-certified in accordance with the
European Directive for in-vitro diagnostic (IVD) medical devices.
Require-
The medac assay satisfies all the requirements for a reliable routine
ments diagnostic test.
- Simple handling - standardised processing and incubation
conditions
- Ready-to-use reagents
- Breakable microtitre strips (individual wells)
- Can be used on automated open microtitre plate systems
- Pipetting control system (indicator system to avoid pipetting
errors)
5medac
EBV EBNA-1 IgG: Short instructions for use
Preparation EBV EBNA-1 IgG ELISA PKS Cat. No. 126-PKS
of the Wash Buffer 1:10 Serum Dilution
reagents 100 ml
900 ml
Wash Water for Sample 1:200
Buffer injections Dilution Buffer
(10x)
Test run Controls, serum samples or plasma,
calibrator (except A1)
Incubation at 37°C,
humid chamber
Wash plate 3 x
200 µl each
Conjugate (except A1)
je 50 µl*
50 µl each 60 min
empty and tap out * when automated 60 µl
Incubation at 37°C, Wash plate 3 x
TMB Substrate Incubation at 37°C,
humid chamber 200 µl each in the dark, humid chamber
50 µl Each
60 min 30 min
empty and tap out
Stop solution (0,5 M H2SO4) Photometric reading
100 µl each O.D.
Reference
450 nm 620 - 650 nm
Calculation Correction of the results:
Nominal OD value of the calibrator x OD measured
ODcorrected = Measured OD of the calibrator
Quantification of the results (under consideration of the batch specific data):
Concentration AU/ml = b / ( OD a
corrected
-1 )
Cut-off = 10 AU/ml.
Grey zone = 9 - 11 AU/ml
Inter- Samples with OD values below the grey zone are reported as
pretation NEGATIVE.
Samples with OD values within the grey zone are reported as
EQUIVOCAL.
Values within the grey zone should be controlled for titer
movement by testing second serum samples after 14 days in
parallel with the initial serum samples.
Samples with OD values exceeding the upper limit of the grey
zone are reported as POSITIVE.
6medac
Contact:
Export
Department: medac GmbH
Diagnostic Division
Cornelia Appelius
Dr. Sabine Dettlaff
Dr. Karen Dreesbach
Theaterstrasse 6
D-22880 Wedel
Tel.: ++49 4103 8006- 0
Fax: ++49 4103 8006 -359
e-mail: c.appelius@medac.de
s.dettlaff@medac.de
k.dreesbach@medac.de
Homepage: www.medac.de
Austria medac
and Dr. Maria Kleindel
Slovenia: Postfach 20
A-3032 Eichgraben
mobile: ++43 676 502 25 69
Fax: ++43 2773 43 574
e-mail: m.kleindel@medac.de
Belgium medac
and Georges Wauthier
Luxembourg: 8, rue des Bailleries
B-5081 Meux
Tel.: ++32 81 74 84 83
Fax: ++32 81 74 84 83
E-mail: g.wauthier@medac.de
Czech medac
Republic Dr. Anna Manthey
and P.O. Box 113
Slovakia: CZ-140 21 Prague 4
Tel.: ++42 02 41 40 26 18
Fax: ++42 02 41 40 26 18
e-mail: a.manthey@medac.de
For all other countries please contact medac Diagnostic Division, Gemany
Edit: 10/05
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