Quantification of de novo central nervous system IgG measles antibody synthesis in SSPE.

De novo central nervous system (CNS) IgG measles antibody synthesis rate was determined in two patients with subacute sclerosing panencephalitis (SSPE). Cerebrospinal fluid and sera were incubated with different concentrations of measles virus to determine maximum absorption of IgG. The difference in IgG concentration before and after incubation was considered the amount of measles-specific IgG antibody. Various tests were done to assess the immunological specificity of the absorption test. Using our empirical formula, the de novo CNS IgG measles antibody synthesis rate for one patient with SSPE was 57 mg/day, or 59% of the total CNS IgG synthesis rate; for the other it was 145 mg/day, or 82%. Daily determination of CNS measles antibody by empirical formula provides a quantification of humoral immunity in SSPE that corrects for both passage of measles antibody across a damaged blood-brain barrier and the effect of an elevated blood antibody level.

Multiple sclerosis serum and cerebrospinal fluid immunoglobulin binding to Fc receptors of oligodendrocytes.

Suspensions of bovine oligodendrocytes were used to study the immunofluorescent staining properties of sera from multiple sclerosis (MS) patients and normal individuals. All sera (14 MS patients, 8 patients with other neurological diseases, and 11 normal individuals) showed positive oligodendrocyte staining by indirect immunofluorescence. Staining persisted after extensive absorption of sera with bovine liver to remove nonspecific binding. Similar findings were obtained for cerebrospinal fluid from all 5 MS patients as well as all 5 patients with other neurological diseases. In additional studies to determine if binding is mediated by the Fc fragment of IgG, results were: (1) ultracentrifuged normal and MS sera failed to react with oligodendrocytes, (2) positive staining with oligodendrocytes was observed after heat aggregation of IgG, (3) ox red blood cells, complexed with antibody, reacted with oligodendrocytes to produce strong rosette formation, and (4) the rosette formation could be blocked by prior treatment of oligodendrocytes with heat-aggregated IgG. The studies fail to support a previous claim for specific binding of immunoglobulins to oligodendrocytes in MS. However, they confirm and extend previous findings that the binding of IgG to oligodendrocytes may not necessarily be an antigen-antibody reaction. Therefore, adherence reactions with a putative antibody must exclude Fc-mediated binding.

Multiple sclerosis de novo CNS IgG synthesis: effect of ACTH and corticosteroids.

ACTH gel and corticosteroids were given to 28 clinically definite multiple sclerosis (MS) patients to determine whether de novo central nervous system (CNS) IgG synthesis (rate and cerebrospinal fluid [CSF] IgG oligoclonal bands) could be eradicated. The most effective treatments were ACTH gel and ACTH gel followed by prednisone, all 11 patients had a significant reduction in rate (p < 0.05), which became normal in eight patients (< 3.3 mg per day). In order of effectiveness, the other drugs used were: dexamethasone or prednisone given orally, and hydrocortisone administered intrathecally. For most treatments, reduction of the rate of CNS IgG synthesis occurred within days and persisted for months after cessation of treatment. The MS CNS immune reaction was not eradicated when IgG synthesis rate became normal, because CSF IgG oligoclonal bands persisted. None of the chronic progressive, severely disabled patients demonstrated significant change in neurologic function or persistent adverse effects.

Multiple sclerosis de novo CNS IgG synthesis. Effect of CNS irradiation.

Megavoltage CNS irradiation was given to 20 patients with clinically definite multiple sclerosis (MS) to determine if de novo CNS IgG synthesis could be eradicated. In all five patients given 1,200 rads, a transient reduction in the de novo CNS IgG synthesis rate was noted. In ten patients given 1,800 rads, the following occurred: a reduction in synthesis rate in three patients, a reduction followed by enhancement in two, only enhancement in four, and no change in one. In all five additional patients, a therapy of adrenocorticotropic hormone (ACTH) followed by prednisone in combination with 1,800 rads produced greater and more persistent decreases in CNS IgG synthesis, but did not block the enhancement effect. Only two of 19 patients who had abnormal CNS IgG synthesis rates had reductions to normal; no patients showed changes in the number or pattern CSF IgG oligoclones. Hence, no treatment eradicated de novo CNS IgG synthesis. A persistent decrease in CSF leukocytes occurred in all 20 patients due to the reduction of small lymphocytes (not dose related). The blood-brain-barrier to albumin concentration was transiently damaged in 11 of 15 patients given irradiation, but when patients were premedicated with ACTH/prednisone therapy, no damage was found. None of the patients demonstrated neurological improvement, change in the activity of their disease, or persistent adverse effects.

Failure of intravenous and intrathecal cytarabine to modify central nervous system IgG synthesis in multiple sclerosis.

To study the nature of the multiple sclerosis (MS) central nervous system (CNS) immune reaction, cytarabine (ara-C) was administered intravenously to three patients and intrathecally to another seven. Although intravenous administration severely suppressed circulating leukocytes derived fom the bone marrow, there was no change in de novo CNS IgG synthesis (rate or presence of CSF IgG oligoclones). Therefore, the MS CNS immune reaction can persist in the presence of severe granulocytopenia and severe monocytopenia of one to two weeks' duration. In four patients who were given ara-C intrathecally, cytotoxic levels (> 10 micrograms/ml) were present for a least 24 hours. A brief elevation in CSF albumin indicated transient damage to the blood-brain barrier. Leukopenia did not result in the seven patients treated intrathecally, and de novo CNS IgG synthesis did not change or was transiently increased. Assuming that cytotoxic levels in CSF diffused to the CNS sites responsible for synthesizing IgG, results indicate that CNS IgG synthesis does not depend on in situ rapid turnover of immune cells. None of the ten patients demonstrated neurological improvement or persistent adverse effects.

Immunofixation electrophoretic techniques applied to identification of proteins in serum and cerebrospinal fluid.

We describe the application of immunofixation staining of agarose-gel electrophoretograms in areas where its use in the clinical laboratory is appropriate. Immunofixation electrophoresis consists of an electrophoretic phase followed by a fixation phase in which antiserum is used to precipitate the protein. As long as the antibody is in slight excess or near equivalency, the antigen/antibody complex remains insoluble. The reaction can be detected by visual inspection in indirect light, by protein staining, or by use of antibodies labeled with fluorescein, enzyme, or isotope. In the method described here we primarily have used protein staining (Coomassie Blue) to accentuate the proteins fixed by antisera. All unreacted proteins are removed by pressing with filter paper and saline washing. In the clinical laboratory, this method expedites immunochemical evaluation of samples and may also supplement immunoelectrophoresis. It has been applied successfully in identifying small obscure monoclonal proteins in the serum and cerebrospinal fluid of patients with multiple sclerosis, subacute sclerosing panencephalitis, biclonal gammopathies, serum monoclonal light chains, and mobility shifts of certain proteins, particularly of the complement series. Immunofixation demonstrates that the protein bands present in spinal fluid from multiple sclerosis and subacute sclerosing panencephalitis patients are of the IgG class of immunoglobulins; and non-IgG protein, such as beta and gamma trace proteins, are not detected. We also comment on reverse immunofixation with labeled antigen as a branch of the procedure that allows detection of function of the immunoglobulins separated by electrophoresis.