Serologic investigations into the etiology of amyotrophic lateral sclerosis

Abstract

The complement fixation, hemagglutination, and passive hemagglutination tests were used to investigate the blood serum of patients with amyotrophic lateral sclerosis, of monkeys with the same disease, and of immunized rabbits for the presence of antibodies. No antibodies could be detected by any of these tests.Previous investigations [1, 2] showed that if rhesus monkeys are infected with the brains of persons dying from amyotr0phic lateral sclerosis (ALS), they develop a disease very similar in its clinical and path,morphological picture to ALS in man.In this investigation an attempt was made to detect specific antibodies in the sera of patients with ALS, of infected monkeys, and of immunized rabbits. No data relating to the discovery of antibodies in chronic infections of the central nervous system could be found in the literature, with the exception of one paper [4] describing very briefly the negative results of a search for antibodies in sheep and goats with scrapie, and giving a list of the various serologic tests used. EXPERIMENTALThe cerebrospinal fluid and brain tissue from patients with, or dying from, ALS and the brain tissue of monkeys infected with the disease and subsequently sacrificed were used as antigens. The brain tissue of persons dying from trauma was used as the control To prepare the antigen the brain tissue was ground with sand in a mortar in the cold for 20 rain, and suspensions made in physiological saline in concentrations of 1 : 5 and 1 : 10 relative to weight of tissue. The suspension was clarified by centrifugation at 2000-4000, 8000, and 13,000 rpm for 10-30 mine The antigen was precipitated and concentrated by centrifugation on a "Spine. ~ ultracentrifuge at 50,000 rpm for 1-2 h~ In addition, V. I~ TovarnitskiUs method, used in 1960 [3] for the diagnosis of tick-borne encephalitis was used to prepare the antigen. For this purpose, the brain tissue was frozen and thawed 5 times, and then precipitated with methyl alcohol. This antigen, in which a certain degree of purification of the brain tissue is achieved by precipitation with methanol, was used in the complement fixation test. The culture fluids used as antigens were obtained by cultivation of a suspension of brain tissue from persons dying from ALS, on human embryonic diploid and kidney ceils. Specimens of tissue culture fluids were obtained at different periods of cultivation of material from patients with ALS, mostly at intervals of 3-4 days. Tissue culture fluids also were obtained during subcultures~ Sera were obtained from patients with ALS, from monkey infected with the disease, and from rabbits immunized with brain suspensions and eerebrospinal fluid. The sera were heated to 56 ~ for 30 rain to destroy thermolabile inhibitors, and diluted with phosphate buffer, pH 7,2. To remove nonspecific agglutinins, the sera were treated with suspensions of the erythrocytes used in the test and a 25% solution of kaolin in borate buffer solution. For the actual test the original suspensions were diluted with borate buffer, pH 9.0, u...