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Isoelectric focusing followed by silver staining. A suitable method for routine investigation of cerebrospinal fluid proteins.

Cerebrospinal fluid (CSF) and serum samples from 361 consecutive patients were investigated. The protein pattern of each sample was examined by isoelectric focusing (IEF) of unconcentrated CSF followed by silver staining and by agar gel electrophoresis of concentrated CSF stained with amido black. 347 paired samples were compared. The results were the same in 83% of the samples. Discordant results were found in 10% of the samples. and in 7% additional information was achieved by one of the methods. IEF was superior to agar gel electrophoresis for the identification of oligoclonal immune reactions of the central nervous system. Tau globulin increase was seen in a few CSF samples, more often with IEF and then associated with other changes of the pattern. IEF of unconcentrated CSF followed by silver staining only requires 20 microliters CSF, while 3-4 ml CSF is needed for the agar gel electrophoretic method. It was concluded that IEF has several advantages compared to agar gel electrophoresis and is suitable as a routine method in clinical laboratories.

Adolescent↗

Estimation of reference intervals for total protein in cerebrospinal fluid.

Protein in cerebrospinal fluid (CSF) was measured by a modified biuret procedure and in two automated instruments, the Du Pont aca and the Kodak Ektachem. The latter's dry-slide reagent was also evaluated for precision, linearity, and effect of potential interferents. In vitro, ampicillin and vancomycin increase the apparent value for CSF protein as measured with the Ektachem slides. We excluded patients with disorders of the central nervous system, and we estimated the central 95% percentile reference intervals for CSF protein for each of the three methods. We found no age or gender dependence of values. By the biuret procedure, the reference interval is 140 to 620 mg/L.

Adult↗

[Cerebrospinal fluid protein levels and blood-cerebrospinal fluid ratio of glucose and electrolytes in infants and children].

The cell and protein content of the cerebrospinal fluid (CSF) and the CSF/serum ratio of glucose, sodium, potassium, calcium, phosphate and chloride were investigated in 71 children without cerebromeningeal illness and aged 0-4 weeks, 5-8 weeks, 3-6 months, 7-12 months, 1-6 years and 7-14 years. The protein content of the CSF was 730 +/- 146 (530-950) mg/l for the age group 0-4 weeks, and 530 +/- 221 (270-950) mg/l for the age group 5-8 weeks. It was significantly higher than for all subsequent age groups. The CSF glucose accounted for 68-82% of the blood glucose. The serum/CSF ratio for sodium ranged from 101 to 104%, for potassium from 55 to 64%, for calcium from 48 to 70%, for phosphate from 24-29%, and for chloride from 106 to 114%. The decrease in serum and CSF potassium and phosphate concentrations from infancy to school age was significant. The greatest difference was evident in the serum/CSF calcium ratio which was significantly higher in the age groups up to 8 weeks than in the subsequent age groups. The electrolyte concentration of CSF with elevated protein and cell values did not differ significantly from CSF of children without cerebromeningeal disorders.

Adolescent↗

Two-dimensional electrophoresis and "ultrasensitive" silver staining of cerebrospinal fluid proteins in neurological diseases.

Cerebrospinal fluid (CSF) proteins, as resolved by two-dimensional electrophoresis and made visible by silver staining, have been examined in patients with various neurological diseases and normal volunteers. The patterns for 15 of 20 patients with Parkinson's disease showed a protein (Mr 25 000) with charge similar to albumin, which was not seen in the patterns for any of 91 normal volunteers. Patterns for 21 of 22 multiple sclerosis patients showed novel immunoglobulin light chain proteins, also not present in the CSF of any normal volunteers. Quantitative analysis by computer-assisted densitometry in Parkinson's disease and multiple sclerosis showed that 20 of 68 and 33 of 85 proteins, respectively, were significantly altered as compared with proteins in the normal population. This ability to characterize both Parkinson's disease and multiple sclerosis molecularly provides a broad baseline for improved clinical diagnosis and may serve as an aid in exploring the underlying pathophysiology. These studies illustrate the potential of applying this methodology in the study of neurological diseases.

Adult↗

The use of high-performance electrophoresis chromatography for the micropurification of cerebrospinal fluid proteins in the rat.

Using reversed-phase high-performance liquid chromatography (HPLC) on a Vydac C8 column in conjunction with sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis (PAGE) and silver staining, we have identified more than 30 proteins in cerebrospinal fluid collected from adult rats by cannulation of cisterna magna. When these partially purified cerebrospinal fluid proteins were further fractionated by high-performance electrophoresis chromatography (HPEC) on an Applied Biosystems 230A HPEC system using a 10% T SDS-polyacrylamide gel with a phosphate base running buffer system under nonreducing conditions, we have purified more than 10 proteins to apparent homogeneity from a pool of 10 ml of rat cerebrospinal fluid as verified by silver staining and direct N-terminal amino acid sequencing. Two additional series of experiments using rat cerebrospinal fluid over a 12-month period yielded virtually identical results. A major advantage of HPEC over conventional HPLC is that the recovery of protein is almost quantitative and is in the range of 90-95% using as little as 1 microgram of protein. The purified proteins from HPEC are ready for direct protein sequencing following a buffer exchange to remove residual Tris and phosphate without additional manipulation. The potential use of HPEC for micropurification of proteins was discussed.

Amino Acid Sequence↗