Counterimmunoelectrophoresis With Serum Prediffusion: An Improved Method for the Detection and Identification of Antibodies Against Extractable Nuclear and Cytoplasmic Antigens

J Immunol Methods. 1997 Feb 14;201(1):89-98. doi: 10.1016/s0022-1759(96)00200-1.

Abstract

In the technique of counterimmunoelectrophoresis (CIE) with serum prediffusion (SPD) serum is allowed to diffuse freely into the gel before pouring the antigenic extract in its trough (or wells) and starting the electrophoresis. Both the immunoprecipitations and the interactions with reference sera are strongly intensified by SPD, leading to higher sensitivity and specificity for the detection of anti-SSA/Ro, anti-SSB/La, anti-U1RNP, anti-Sm, anti-Jo1 and even anti-Scl-70 antibodies. We found that the optimal SPD time was 2 h. To evaluate the relevance of SPD for the clinical laboratory, 92 antinuclear antibody (ANA) positive sera were tested on CIE without SPD and with 2 h SPD in identification tests with SSA/Ro, SSB/La, Sm, U1RNP and Jo1 reference sera (rsa). The precipitation lines and their interactions were evaluated by three independent observers. It was observed that SPD considerably improved the efficiency of CIE for antibody identification. The mechanisms underlying the intensification of the precipitation lines by SPD are discussed as are the characteristics of the CIE in comparison with other test systems such as the enzyme linked immunosorbent assay (ELISA) and immunoblot.

MeSH terms

  • Animals
  • Antibodies, Antinuclear / analysis*
  • Autoantibodies / analysis*
  • Autoantigens / immunology
  • Blood
  • Counterimmunoelectrophoresis / methods*
  • Cytoplasm / immunology*
  • Diffusion
  • Humans
  • Immunodiffusion / methods
  • Nuclear Proteins / immunology*
  • Precipitin Tests
  • Rabbits

Substances

  • Antibodies, Antinuclear
  • Autoantibodies
  • Autoantigens
  • Nuclear Proteins