Chemiluminescent detection of oligoclonal immunoglobulins after isoelectric focusing and affinity-mediated immunoblotting

Abstract

Background and Aims: Detection of oligoclonal IgG (o-IgG) in the cerebrospinal fluid (CSF) not found in serum is the principal laboratory test to support a diagnosis of multiple sclerosis. The aim of this study was to compare chemiluminescent and chromogenic detection of oligoclonal immunoglobulins in the cerebrospinal fluid and serum after their separation by means of isoelectric focusing followed by immunoblotting. Methods: A set of experiments was designed to detect oligoclonal immunoglobulins by means of alkaline phosphatase BCIP/NBT substrate and chemiluminescent peroxidase substrate. Results: Based on visual evaluation of signals, chemiluminescent detection requires about a 4 times lower amount of applied protein than very sensitive BCIP/NBT chromogenic detection. Very good correlation between methods has been shown for oligoclonal IgG. Antigen-specific oligoclonal IgG could be demonstrated by both methods although the pattern was clearer using chemiluminescence. In one patient, oligoclonal IgD bands barely visible by BCIP/NBT were convincingly demonstrated by chemiluminescence. Conclusion: Chemiluminescent detection is a feasible option for oligoclonal immunoglobulin detection and could be used in cases when the sensitivity needs to be improved. Further studies and method optimisation are warranted.

Keywords

• cerebrospinal fluid
• chemiluminescence
• immunoglobulins
• isoelectric focusing
• immunoblotting