Line immunoassay for parallel detection of 9 different autoantibodies in the serological differential diagnosis of PBC
A. Janssen1, L. Komorowski1, D. Bogdanos2, C. Probst1, W. Meyer1,
T. Scheper1, W. Schlumberger1, and W. Stoecker1
1
Institute for Experimental Immunology, affiliated to EUROIMMUN AG, Luebeck, Germany
Institute Of Liver Studies, King’s College London School Of Medicine, King’s College Hospital, London, UK
Sp100 and/or
PML pos.
l
gp210 pos.
-5
nt
2
ro
P
4
Co
Ro
LC
SL
A
-1
/L
-1
M
0
LK
21
gp
L
PM
A
M
A
-M
2
M
(s 2-3
yn E
on
ym
Sp
:B
10
PO
0
)
2
ALD
PBC
ALD
PBC
22
4
1
13
27
neg./neg./neg.
11
88
ALD
AIH
ALD
AIH
ALD
HBD
AMA-M2 and/or
M2-3E (BPO) pos.
Prevalence of antibodies in 170 PBC patients
Introduction
Primary biliary liver cirrhosis (PBC) is associated with different serum autoantibodies
against antigens found in mitochondria, nuclear dots and nuclear membrane. Analysis
of these antibodies helps to discriminate
between PBC and autoimmune hepatitis
(AIH). We evaluated a new robust multiparametric test system for diagnosing autoimmune liver diseases.
Methods
A line immunoassay was created using
native full-size AMA-M2 and the following
recombinant proteins: M2-3E (synonym:
BPO; a fusion protein of the immunogenic
lipoyl domains of branched-chain oxoacid
dehydrogenase, pyruvate dehydrogenase
and oxoglutarate dehydrogenase)1, Sp100
(spot-pattern 100kDa protein) and PML
(promyelocytic leukaemia protein), both
representing antigens from nuclear dots,
gp210 (glycoprotein 210, part of the nuclear pore complex), LKM-1 (liver-kidney
microsomes), LC-1 (cytosolic liver antigen
type 1), SLA/LP (soluble liver antigen/liverpancreas antigen) and Ro-52 (anti-Ro-52
antibodies in combination with anti-SLA/LP
are discussed as potential markers for an
unfavourable disease course).
This profile was used to screen for antibodies in sera of 170 patients with clinically
characterised PBC, 49 with AIH, 200 with
viral hepatitis (HBV or HCV) and 50 healthy
blood donors. Prevalence and intensity of
the bands were automatically evaluated
using a commercial computer programme
(EUROLineScan, EUROIMMUN, Germany).
Results
In 94% of the PBC sera, antibodies against at
least one of the antigens AMA-M2, M2-3E,
Sp100, PML, gp210 were detected with a
specificity of 99% as referred to the panels of
viral hepatitis and blood donors. Four out of
six positive samples in the AIH group could
be attributed to PBC/AIH overlap patients after re-evaluation of the medical records.
Discussion
Using the new comprehensive profile, a
serological diagnosis of PBC can be made
with a yet unequalled sensitivity of 94%.
Autoantibodies against the designer antigen M2-3E and native AMA-M2 play a predominant role in the majority of patients,
with M2-3E contributing the most to the
sensitivity of the assay. However, in 6% of
the cases immune reactivities were exclusively directed against Sp100, PML and/or
gp210. These patients would not have been
detected by immunoassays using AMA-M2
as the single antigen. In sum, the novel
line immunoassay represents an outstanding diagnostic tool for PBC. It exceeds all
previous detection methods with regard to
sensitivity and specificity, including testing
by indirect immunofluorescence.
Autoantibody prevalence
Collective
n
AMA-M2
M2-3E
(BPO)
AMA-M2
and/or
M2-3E
Sp100
PML
Sp100 and/
or PML
gp210
At least
one of all
PBC
170
138
(81%)
146
(86%)
150
(88%)
35
(21%)
22
(13%)
40
(24%)
45
(26%)
159
(94%)
AIH
49
4
(8%)
2
(4%)
4
(8%)
2
(4%)
2
(4%)
2
(4%)
2
(4%)
6
(12%)
HBV or HCV
200
0
0
0
0
1
(0.5%)
1
(0.5%)
1
(0.5%)
2
(1%)
Blood donors
50
0
0
0
0
0
0
0
0
Literature: 1Moteki et al. Use of a designer triple expression hybrid clone for three different lipoyl domain for the detection of antimitochondrial autoantibodies. Hepatology. July 1996, 24 (1): 97-103.
Scientific presentation at the 10th International Workshop on Autoantibodies and Autoimmunity (Guadalajara, Mexico, March 2008)
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