A major Sm epitope anchored to sequential oligopeptide carriers is a suitable antigenic substrate to detect anti-Sm antibodies.
A sensitive, highly reproducible, solid-phase enzyme immunoassay (ELISA), was developed in order to investigate whether the synthetic heptapeptide PPGMRPP-a major epitope of the Sm autoantigen-anchored in five copies to a sequential oligopeptide carrier (SOC), [(PPGMRPP)5-SOC5] is a suitable antigenic substrate to identify anti-Sm/antibodies. Sera with different autoantibody specificities [45 anti-Sm, 40 anti-U1RNP, 40 anti-Ro (SSA)/La(SSB) positive, 21 Antinuclear antibody positive, but negative for antibodies to extractable nuclear antigens (ANA + /ENA - ) and 75 normal human sera, ANA negative] and 75 sera from patients with rheumatoid arthritis (RA) were tested for anti-(PPGMRPP)5-(SOC)5 reactivity in order to evaluate the specificity and sensitivity of the method to detect anti-Sm antibodies. RNA immunoprecipitation assays for the detection of anti-Sm and anti-U1RNP antibodies and counter immunoelectrophoresis (CIE) for the detection of anti-Ro(SSA) and anti-La(SSB) antibodies were used as reference techniques. The sensitivity of the method was 98% and the specificity was 68% for the determination of anti-Sm antibodies, while for the determination of anti-Sm and/or anti-U1RNP reactivity (antibodies to snRNPs) the corresponding values were 82% and 86%, respectively. In a comparison of the above assay with an ELISA, using Sm/U1RNP purified complex as immobilized antigen it was shown that the sensitivity of the anti-Sm/U1RNP ELISA in detecting anti-snRNPs was 74%; in addition sera with anti-Sm antibodies gave higher binding in the anti-(PPGMRPP)5-(SOC)5 ELISA compared with anti-Sm/U1RNP ELISA. Intra- and inter-assay precision was measured on four sera with reactivities extending into a wide range of absorbance values showed that the intra-assay coefficient of variation (CV%) ranged from 2.7 to 6 and the inter-assay CV% ranged from 9 to 14.5. These results indicate that the PPGMRPP peptide anchored to a pentameric SOC as a carrier is a suitable antigen for detecting anti-Sm antibodies and that the above ELISA is a rapid, reproducible and valuable screening method to test anti-Sm/U1RNP reactivities.
Petrovas
C J
CJ
Department of Pathophysiology, Medical School, National University of Athens, Greece.
Vlachoyiannopoulos
P G
PG
Tzioufas
A G
AG
Alexopoulos
C
C
Tsikaris
V
V
Sakarellos-Daitsiotis
M
M
Sakarellos
C
C
Moutsopoulos
H M
HM
D000974:Antibodies, Antinuclear / Q000276:immunology* / Q000302:isolation & purification; D001324:Autoantigens / Q000276:immunology*; D001667:Binding, Competitive; D004337:Drug Carriers; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000276:immunology*; D006801:Humans; D009842:Oligopeptides; D015203:Reproducibility of Results; D017411:Ribonucleoproteins, Small Nuclear; D012680:Sensitivity and Specificity; D055517:snRNP Core Proteins
D000974:Antibodies, Antinuclear; D001324:Autoantigens; D004337:Drug Carriers; D000939:Epitopes; D009842:Oligopeptides; D017411:Ribonucleoproteins, Small Nuclear; D055517:snRNP Core Proteins