Immobilized pH gradient isoelectric focusing of cerebrospinal fluid proteins.
ABSTRACT Isoelectric focusing in immobilized pH gradients, supplemented with 0.5% w/v carrier ampholytes was applied for studies of native proteins, especially immunoglobulin G, in cerebrospinal fluid and serum. All 72 paired samples were run on pH 4-10 gels; 25 of them were also examined in pH 7-10 gels. Silver staining and nitrocellulose blotting with amplified immunoperoxidase detection of immunoglobulin G were used for protein visualization. Intrathecally produced immunoglobulin G was resolved into sharply focused, straight and easily identifiable fractions. The pH gradients were stable and the inter-gel reproducibilities of individual immunoglobulin G patterns were good.
- Scandinavian Journal of Clinical and Laboratory Investigation 10/1977; 37(5):385-90. · 1.29 Impact Factor
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ABSTRACT: To demonstrate oligoclonal IgG bands (I) in unconcentrated cerebrospinal fluid, we used isoelectric focusing in agarose followed by protein transfer to cellulose nitrate membrane, double-antibody peroxidase labeling, and avidin-biotin amplification. I can be reliably seen after isoelectric focusing of 5-microL specimens containing 125 ng of IgG (25 mg/L). Thus the technique is more sensitive than others (e.g., silver staining) and more reliable than radioimmunofixation. When we used this technique with fluids from 62 patients with multiple sclerosis and infectious disease of the central nervous system, 84% displayed I, a percentage not increased when the same specimens were concentrated to 3.5 g of IgG per liter, examined by agarose isoelectric focusing, and stained with Coomassie Blue. Results for 53 patients with tension headache and psychoneurosis were all negative. By obviating the need to concentrate samples of cerebrospinal fluid the present method is a useful, sensitive alternative for demonstrating I.Clinical Chemistry 08/1984; 30(7):1246-9. · 7.15 Impact Factor
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ABSTRACT: Unconcentrated cerebrospinal fluid (CSF) and serum samples from 1114 consecutive patients were examined for presence of oligoclonal IgG bands (OB) by agarose isoelectric focusing (AIF) followed by protein transfer to nitrocellulose membrane, immunolabeling, and avidinbiotin-peroxidase staining (avidin-biotin AIF). Oligoclonal bands were demonstrated in CSF from all 58 patients with multiple sclerosis (MS), eight of 29 with aseptic nervous system infections, and 9% of 1014 with other neurological disorders (OND) considered as noninflammatory at primary clinical evaluation. Comparative examination of all specimens in another laboratory by conventional AIF after concentration of CSF revealed lower frequencies of OB in all diagnostic groups. In addition to the high sensitivity of avidinbiotin AIF, which enables detection of OB by separation of 5 microL of unconcentrated CSF even when the CSF IgG level is around the lower normal range, the procedure also has optimal specificity since IgG exclusively is detected. Avidin-biotin AIF may be the method preferred for routine examination of CSF for OB. Demonstration of OB in CSF is valuable especially in MS, where, in contrast to diagnostic aids such as evoked potentials and neuro-imaging, it establishes inflammatory type of nervous system involvements. Oligoclonal IgG bands in CSF from patients with OND reflect intrathecal immune response and should lead to investigations of infectious etiology.JAMA Neurology 11/1987; 44(10):1041-4. · 7.58 Impact Factor