MAT-1, a Monoclonal Antibody that Specifically Recognizes Human Tyrosinase

Abstract

There is no available monoclonal antibody which reacts specifically recognizes human tyrosinase. Employing a synthetic peptide, MEKEDYHSLYQSHL, corresponding to the carboxyl terminus of human tyrosinase as an immunogen, we produced a mouse monoclonal antibody MAT-1 of the IgG1 isotype. The epitope for MAT-1 was determined to be EDYH, the sequence of which is not present in human tyrosinase-related protein-1 (TRP-1) or tyrosinase-related protein-2 (TRP-2). By transient expression assays and immunofluorescence technique, we show that MAT-1 reacts specifically with cells expressing human tyrosinase cDNA but not with cells expressing TRP-1 or TRP-2 cDNA. The results of immunohistochemical staining also confirmed that MAT-1 reacts specifically with epidermal melanocytes in human skin sections. MAT-1 should be invaluable for studying the interaction between tyrosinase and TRPs and for detecting the changes in the levels of tyrosinase expression. In addition, MAT-1 should be useful as a sensitive immunohistochemical tool for investigation of various pigmentary disorders and possibly for the diagnosis of melanoma.

Detection of Mouse Tyrosinase With a Monoclonal Antibody MAT‐1 Against Human Tyrosinase

“…We successfully prepared the anti-human tyrosinase monoclonal antibody (MAT-l), which reacts specifically with Dulbecco's modified eagle's medium (DMEM) were purchased from Gibe0 BRL, Tokyo, Japan; fetal bovine Serum (FBS) from Whittaker, Walkersville, MD; GIT medium and L-P-(3,4-dihydroxyphenyl) alanine (DOPA) from Wako Pure Chemical Industries, Ltd., Osaka, Japan. human tyrosinase but not with human TRP-1 and TRP-2 (Takimoto et al, 1995). The epitope for MAT-1 is comprised of four amino acids (EDYH sequence); MAT-1 also reacts with the amino acid sequence of DYH to a lesser extent (Takimoto et al, 1995).…”

“…human tyrosinase but not with human TRP-1 and TRP-2 (Takimoto et al, 1995). The epitope for MAT-1 is comprised of four amino acids (EDYH sequence); MAT-1 also reacts with the amino acid sequence of DYH to a lesser extent (Takimoto et al, 1995). Incidentally, mouse tyrosinase has the DYH sequence (Yamamoto et al, 1987) at the position corresponding to the EDYH sequence of human tyrosinase (Shibahara et al, 1988).…”

“…Monoclonal Antibody (MAT-1) The hybridoma cells that produce anti-human tyrosinase monoclonal antibody (MAT-1) were cultured in GIT medium at 37°C under 5% COZ (Takimoto et al, 1995). The MAT-1 was purified from the conditioned medium by ammonium sulfate precipitation, followed by further purifica-…”

Construction of pigmented skin equivalent and its application to the study of congenital disorders of pigmentation

“…Pigmented skin equivalents were mounted in blocks, cut into 4-mm vertical sections, and stained with haematoxylin and eosin. For immunohistochemical staining, mouse monoclonal antibodies to human tyrosinase (MAT-1) [15] were purchased from Pola R&D Laboratories (Yokohama, Japan). The sections were lightly counterstained with haematoxylin, mounted, and looked at with a light-microscope (Nikon Microphoto-FXA).…”

Expression of Tyrosinase-related Protein 2/DOPAchrome Tautomerase in the Retinoblastoma

“…The cDNA probe for TRP-2/DCT was the SacI/EcoRV fragment derived from human TRP-2 cDNA (Yokoyama et al, 1994a). The full-length cDNA probes for human tyrosinase and TRP-1 were the SalI-XbaI fragment derived from pRHOHT2 (Takeda et al, 1989) and the SalI-XbaI fragment derived from pRHOHTa (Takimoto et al, 1995), respectively. Expression of glyceraldehyde-3-phosphate dehydrogenase (G3PDH) mRNA was examined as an internal control.…”