1. A fast and sensitive method for measuring picomole levels of total free amino acids in very small amounts of biological tissues.
- Author
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Fisher GH, Arias I, Quesada I, D'Aniello S, Errico F, Di Fiore MM, and D'Aniello A
- Subjects
- Amino Acids chemistry, Animals, Ganglia, Invertebrate chemistry, Ganglia, Invertebrate metabolism, Mercaptoethanol chemistry, Ovum chemistry, Ovum metabolism, Pineal Gland chemistry, Pineal Gland metabolism, Pituitary Gland chemistry, Pituitary Gland metabolism, Rats, Sensitivity and Specificity, Spectrometry, Fluorescence, gamma-Aminobutyric Acid analysis, gamma-Aminobutyric Acid chemistry, o-Phthalaldehyde chemistry, Amino Acids analysis, Biochemistry methods
- Abstract
In the present study we describe a simple and fast method to measure the concentration of total free amino acids in very small amounts of biological tissues. The procedure described here is based on the reaction of free amino acids with o-phthaldialdehyde (OPA) in the presence of a reducing agent, beta-mercaptoethanol (MET), to give a complex which can be measured by fluorescence. It is a very rapid process and has the same reliability as the conventional ninhydrin method of Moore and Stein but is about 500 times more sensitive. The sensitivity of the new protocol is such to permit the determination with high reliability of very small amounts of free amino acids at picomole levels, either in a standard amino acid mixture or in biological tissues, without chromatographic separation of the amino acids. It is particularly useful when the amount of the sample is very low, e.g. on a single pituitary or pineal gland of small animals or on single cells, such as oocytes or eggs, as well as single ganglions or axons of marine invertebrates.
- Published
- 2001
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