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A Histochemical Method for the Demonstration of Leucocyte Naphthylamidase Using Two Substrates: Alanyl‐ and Methionyl‐ 4 ‐Methoxy‐ 2 ‐Naphthylamide
Author(s) -
Rutenburg A. M.,
Rosales C. L.
Publication year - 1966
Publication title -
british journal of haematology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.907
H-Index - 186
eISSN - 1365-2141
pISSN - 0007-1048
DOI - 10.1111/j.1365-2141.1966.tb00142.x
Subject(s) - substrate (aquarium) , enzyme , incubation , hydrolysis , biochemistry , yield (engineering) , chemistry , biology , materials science , ecology , metallurgy
H istochemical methods for the demonstration of naphthylamidase* in tissues were based on the coupling properties of 2‐naphthylamine (Burstone and Folk, 1956; Nachlas, Crawford and Seligman, 1957) after enzymatic release from amino acid naphthylamides. But significant diffusion of the amine and the lack of a suitable diazonium salt to achieve a sufficiently prompt capture reaction to form an insoluble dye as well as prolonged incubation prevented adequate histochemical localization. When the coupling rate of 2‐naphthylamine was found to be significantly enhanced by the presence of a methoxy group in the 4‐position (Nachlas, Goldstein, Rosenblatt, Kirsch and Seligman, 1959), l ‐leucyl‐ 4 ‐methoxy‐ 2 ‐naphthylamide (Rosenblatt, Nachlas and Seligman, 1958) was synthesized and found to serve as a more satisfactory substrate for naphthylamidase demonstration (Nachlas, Morris, Rosenblatt and Seligman, 1960). Replacement of the leucyl group in leucyl‐ 2 ‐naphthylamide by alanyl or methionyl groups resulted in a more rapid hydrolysis by naphthylamidase (Nachlas, Goldstein and Seligman, 1962; Smith, Kaufman and Rutenburg, 1965; Monis, Wasserkrug and Seligman, 1965). It therefore appeared likely that the substrates alanyl‐ and methionyl‐ 4 ‐methoxy‐ 2 ‐naphthylamides would yield further improvement of localization of enzymatic sites and would be potentially useful for the demonstration of naphthylamidase in tissues such as white cells which are known to have a low content of this enzyme. In this report we present the data on the use of both substrates for the demonstration of naphthylamidase in human leucocytes.