Abstract
Most patients with multiple sclerosis synthesize an abnormal protein known as oligoclonal IgG (intrathecal synthesis) and its detection in cerebrospinal fluid is important in the diagnosis of multiple sclerosis. Some is more alkaline than normal IgG which helps in its identification. The highest detection rates are obtained by isoelectric focusing with carrier ampholytes to form the pH gradient. Carrier ampholyte pH gradients are not stable, especially at high pH values so that very alkaline proteins, such as intrathecal IgG, are not well resolved. A new isoelectric focusing method has been developed for this research that uses an immobilised pH gradient in polyacrylamide gel over the pH range 7-10 which produces a linear and stable pH gradient with excellent resolution. It allows highly alkaline proteins to be separated and studied in more detail than before. After isoelectric focusing, the protein patterns were transferred onto polyvinylinedifluoride membranes and visualised using immuno-enzyme detection methods. Only 50 patients were investigated but highly alkaline intrathecal IgG bands were found in the cerebrospinal fluid of all the multiple sclerosis patients (n=14). The specificity of this method for the diagnosis of multiple sclerosis was 98%. Studies on immunoglobulin light chains of the oligoclonal IgG bands were difficult because of antiserum crossreactivity. No free immunoglobulin light chains were detected, between pH 7-10, in the 19 CSF samples analysed. The method also lends itself to studies of the properties that lead to the observation of alkaline IgG bands in multiple sclerosis. Some such studies were performed. Thus, the addition of myelin basic protein to IgG indicated binding between these proteins which may be a factor in the formation of highly alkaline IgG. Most of the alkaline oligoclonal IgG bands seem to exist with low carbohydrate (sialic acid) content and this property may give rise to their characteristic high isoelectric points.