[Agarose gel isoelectric focusing: application to the study of abnormalities of immunoglobulin clonality in CSF and serum].
Since it is quite difficult to commonly use isoelectric focusing (IEF) of proteins in polyacrylamide gel for biological diagnosis, we have developed a method based on IEF in agarose gel, to split proteins from sera and cerebrospinal fluid (CSF). A prefocalisation at low voltage (250 V) is made on a custom thin gel of agarose (0.5 mm) containing some carrier ampholytes (pH 5-9). After deposition of biological samples, the gel is run at 500 V, thereafter at 1200 V. After focusing, the gel is fixed before being coloured by a simplified silver staining technique. In order to demonstrate the good resolution of the immunoglobulines (Ig) in the pH gradient, a transfer on a nitrocellulose membrane followed by an immunofixation was carried out from unstained gels after IEF. This separation on agarose gel shows several advantages, ie its speed (3H total), its lack of toxicity, its sensibility and its reproductibility. It is specially well suited for the diagnosis of diseases characterised by oligoclonal or monoclonal Ig, particularly those found in the CSF during neurologic diseases like multiple sclerosis. Several examples of focused sera and CSF are reviewed in the paper.
Lebrun-Fourcy
C
C
Laboratoire de biochimie A, CHU de Grenoble, France.
Rondot
J
J
Revol
C
C
Renversez
J C
JC
D003001:Cloning, Molecular; D004587:Electrophoresis, Agar Gel / Q000379:methods*; D007136:Immunoglobulins / Q000097:blood* / Q000134:cerebrospinal fluid* / Q000378:metabolism; D066298:In Vitro Techniques; D007525:Isoelectric Focusing / Q000379:methods*; D009103:Multiple Sclerosis / Q000097:blood / Q000134:cerebrospinal fluid
D007136:Immunoglobulins
L'isoélectrofocalisation sur gel d'agarose: application a l'étude des anomalies de clonalité des immunoglobulines dans le LCR et le sérum.