Improved Procedure for Screening Expression Libraries for Novel Autoantigens

Document Type: Research Paper

Authors

1 National Research Center for Genetic Engineering and Biotechnology (NRCGEB),

2 Biotechnology Research Group, Murdoch University,

Abstract

The standard method for immunoscreening of a cDNA expression library is time-consuming because
of the production of a large proportion of false positives during the first and second round of screening.
This problem is more important when a sensitive chemiluminescence detection system is used. Due to
the high sensitivity of the detection system, there is a need to avoid false positives which occur when the
antibody reacts non-specifically. False positives are generally eliminated through absorption of the antibody
with the host bacteria and by eliminating any clones, which react with antibodies present in normal
sera. Here we present a method of obtaining almost identical bacteriophage plates by culturing phage in
parallel, and show that this technique produces positive plaques in duplicate and eliminates false positives.
Using this method, we successfully screened a human fetal spinal cord lambda gt11 cDNA library
using purified immunoglobulin G (IgG) from patients with multiple sclerosis (MS) and Guillain – Barre syndrome (GBS).

Keywords