Human 14‐3‐3 Protein: Radioimmunoassay, Tissue Distribution, and Cerebrospinal Fluid Levels in Patients with Neurological Disorders

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Abstract
An antiserum to human 14‐3‐3 protein has been produced in rabbits. The protein was a poor antigen and attempts to improve immunogenicity were unsuccessful. A radioimmunoassay was developed using the antiserum, 125I‐ 14‐3‐3‐2, and unlabelled 14‐3‐3‐2 as standards. The assay had a sensitivity limit of 2.5 ng.m1−1. The minor component of human 14‐3‐3 protein (14‐3‐3‐1 protein) cross‐reacted to approximately 10% in the assay. Human tissues were surveyed for 14‐3‐3 protein by two‐dimensional electrophoresis and by radioimmunoassay. Two‐dimensional electrophoresis showed a 14‐3‐3 protein complex in brain, intestine, and testis, but not in other tissues. Radioimmunoassay showed that although brain had the highest concentration of 14‐3‐3 (13.3 μg. mg−1 soluble protein), immunoreactivity was present in all tissues, with the concentration in intestine and testis approaching 50% of the brain level. Lower levels (less than 1.0 μg. mg−1 soluble protein) were seen in liver, kidney, skeletal muscle, and erythrocytes. The immunoreactivity present in tissues other than brain showed the same molecular weight and charge characteristics as authentic 14‐3‐3 protein. The radioimmunoassay also detected 14‐3‐3 protein in serum (50 ng.m1−1) and in CSF (5‐130 ng.ml−1). The immunoreactivity present in CSF appeared to be intact 14‐3‐3 protein. CSF 14‐3‐3 levels were measured in 82 patients with various neurological disorders. Measurements of this protein did not appear sufficiently discriminating to be o f diagnostic value.

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