This study evaluated antinuclear antibodies (ANAs) in 600 patient serum samples using multiple techniques including immunofluorescence (IIF), AtheNA Multi-lyte, and Bioplex 2200. Bioplex 2200 detected 33 samples positive for anti-SSA/Ro52 antibodies, 42 for anti-SSA/Ro60, and 17 for anti-La/SS-B. There was 96.5-98.3% concordance between Bioplex 2200 and AtheNA Multi-lyte. IIF detected 48 positive samples, with various staining patterns. The study concludes that Bioplex 2200 is a good option for multiplex detection of autoantibodies associated with diseases like

1. EVALUATION OF ANTINUCLEAR RO/SS-A AND LA/SS-B
ANTIBODIES BY MULTIPLEX TECHNOLOGY AT THE UNIVERSITY
HOSPITAL “12 DE OCTUBRE” (MADRID-SPAIN)
M. Talise1, V. Peréz1, M. Sevilla 1, S. Lermo1, G. Badra2, L. Borque3, A. Serrano1
1Immunology Service, University Hospital 12 de Octubre, 2University Rey Juan Carlos, Madrid, 3San Pedro Hospital, Logroño, Spain
INTRODUCTION
Autoantibodies to Ro/SS-A and La/SS-B ribonucleoproteins are found 45
in autoimmune diseases such as primary Sjögren´s syndrome, 40
systemic lupus erythematousus (SLE) and rheumatoid arthritis1 (RA).
35
Two types of anti-Ro/SSA antibodies have been described, anti-
30
SSA-52 kDa and anti-SSA-60kDa, historically, these autoantibodies
were considered as a uniform autoantibody-system. However, recent 25
studies provided evidence that Ro60 and Ro52 are not part of a 20
stable macromolecular complex and that anti-Ro52 and anti-Ro60 15
(SS-A) antibodies have different clinical associations and each
10
specific to different antigens2, for this reason it is necessary to be
5
able to individually identify these antibodies in the autoimmunity
laboratory and the Bioplex TM 2200 automated system offers this 0
Anti-SSA Athena Anti-SSA Bioplex Anti-SSA/Ro52 Anti-SSA/Ro60 Anti-SSA Ro52 and Anti-SSB/La Anti-SSB/La
2200 Bioplex Bioplex Ro60 Bioplex Athena Bioplex 2200
advantage; we compare this technique with the immunofluorescence
(IIF) and AtheNA Multi-lyte® ANA test system a multiplex fluorescent We observed 15 discrepancies: Anti-SSA negative by Athena but
microsphere immunoassay for the semi-quantitative of Ig G class positive Anti-SSA/Ro52 and/or Anti-SSA/Ro60 by Bioplex 2200; of
antibody to 9 separate analytes (DNA, SSA, SSB, Sm, RNP, Scl-70, which 12 were IIF positive and had some autoimmunity disease: 6
Jo-1, Centromere B, and Histone). SLE; 2 Systemic Sclerosis, 2 autoimmune Hepatitis, 1 Sjögren´s
syndrome + Raynaud , 1 primary biliary cirrhosis, 1
METHODOLOGY dermatomyositis. Only three discrepancies were Anti-SSA Athena
We studied sera from 600 patients randomly admitted to the positive, Anti-SSA Bioplex Negative and IIF negative, one had
autoimmunity section during May and June 2009, these sera were rheumatoid arthritis.
obtained from patients referred from primary care physicians, general On the other hand other 8 discrepancies were observed: Anti-
internists, rheumatologists and other subspecialties such as SSB Athena’s Negative but Bioplex 2200 Positive, of which 8 were
nephrology, dermatology, etc . Antinuclear antibodies (ANA´s ) were IIF positive and four had diagnosis of SLE, 2 RA, 2 Sjögren´s Sx.
determined by immunofluorescence (IIF) on HEp-2 cell, AtheNA Multi- Only two sera were Anti-SSB Athena positive but Bioplex 2200
lyte® multiplex technique and BioPlex™2200 (by Bio-Rad Laboratories, negative, of which 1 was IIF positive and 1 had RA.
Hercules, CA) a fully automated Luminex based system developed for
CONCLUSIONS
analysis of 13 antibodies including anti-SSA-52 kDa and anti-SSA-60
We conclude that BioPlex2200 offers a good percentage of
kDa in a primary tube and SS-B, Sm, Sm/RNP, RNPA, RNP-68, Scl70,
concordance with other methods, and it can be a good solution as
C e n t r o m e r e - B , d s D N A , c h r o m a t i n , J o 1 , r i b o s o m a l P.
a multiplex approach to diagnose autoimmune diseases like
Immunofluorescence was considered positive from the dilution 1/ 80.
Sjögren´s syndrome , systemic lupus erythematousus and
RESULTS rheumatoid arthritis because the separation of Ro / SSA 52 and 60
Six hundred sera were evaluated, 33 sera were Anti-SSA/Ro52, 42 Anti- antigens conferred increased sensitivity, provides greater
SSA/Ro60, and 17 Anti-La/SS-B positive by Bioplex 2200; 37 sera were specificity with a lower rate of false negatives.
positive to Anti SS-A/Ro and 11 AntiSS-B/La by AtheNA Multi-lyte® with a
REFERENCES
global concordance of 96.5% and 98.33% respectively; by IIF we obtained 1. van Woerkom JM, Geertzema JG, Nikkels PG, et al. Expression of Ro/SS-A and La/
48 positive sera, 25 (52.08%) with coarse and fine speckled pattern, 18 SS-B determined by immunohistochemistry in healthy, inflamed and autoimmune
diseased human tissues: a generalized phenomenon. Clin Exp Rheumatol. 2004 May-
(37.50%) homogeneous, 2 (4.20%) Centromere-B, 2 (4,20%) Nuclear dots.
Jun;22(3):285-92.
Finally, 80.70% of the patients had at least one diagnosis of autoimmune
2. J. Schulte-Pelkum a,M. Fritzler , M. Mahler . Latest update on the Ro/SS-A
disease. autoantibody system. Autoimmunity Reviews 8 (2009) 632–637