Automated Fluorometric Assays

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Abstract

The published automated fluorometric histamine assays are based approximately on the original procedure described by Shore et al. (1959) (Ruff et al. 1967, 1977; Evans et al. 1973; Martin and Harrison 1973; Siraganian 1974, 1975, 1976; Parsons et al. 1980; Reinders et al. 1980; Wilhelms 1980; Assem and Chong 1982; Lebel 1983) and with one exception (Stahl Skov et al. 1984) run with continuous flow systems, usually purchased from Technicon. The system as described and developed by Stahl Skov et al. (1984) will be described in Sect. IX. An automated sampler is used to dispense the reference histamine standard solutions, the wash reagent (usually water or perhcloric acid) and the samples into the system. Precise volumes of the stream components (samples, chemicals, air) are delivered into the system by a proportioning pump with calibrated pump tubes. In the analytical cartridge the stream components are mixed in the correct sequence (chemical reaction); thereafter the fluoresence is measured with a fluorometer and recorded on a chart recorder. The methods all have the major advantage that they are quick to perform (12–60 samples/h), use small sample volumes (ca. 0.5–1 ml) and, although fairly expensive to purchase, are not labour intensive. They have usually been applied to situations where a large number of samples are produced containing relatively high histamine concentrations (>10 ng/ml), e.g. histamine release from isolated mast cell suspensions, tissue histamine.