Tetrazoliummethoden zum histochemischen Hydrolasennachweis

Abstract

Using freeze-dried or sections from fresh-frozen or aldehyde-fixed material nitro BT (NBT), tetranito BT (TNBT), distyryl nitro BT (DS-NBT), thiocarbamyl nitro BT (TC-NBT) or benzothiazolylstyrylphthalhydrazidyl tetrazolium chloride (BSPT) were tested as auxiliary reagents for the localization of glycosidases, phosphatases and non-specific esterases with indoxyl substrates in rat tissues. By means of NBT or TNBT as a tetrazolium salt acid beta-D-galactosidase, beta-D-N-acetylglucosaminidase, acid phosphatase, neuraminidase and non-specific esterase can only be localized at the cellular level; a more precise localization is possible for lactase-beta-D-glucosidase in the intestinal brush border, and the best results are obtained in the demonstration of alkaline phosphatase; among all methods described previously the tetrazolium procedure with TNBT is the method of choice for the light microscopic localization of this enzyme. Reverse data are observed with BSPT as a tetrazolium salt; then, all acid and neutral hydrolases can be exactly localized in lysosomes, secretion granules, cytoplasm and/or microvilli of many cells and tissues provided BSPT-formazan is stabilized by osmification. Furthermore, this procedure enables the reliable ultracytochemical demonstration of these enzymes. However, in the case of alkaline phosphatase only sites with high enzyme activity reveal a positive reaction. -DS- and TC-NBT are inferior to NBT, TNBT or BSPT.

Enzyme Histochemistry

“…The ever-present danger of overoxidation with ferri-ferrocyanide can be avoided by the use of phenazine methosulfate (PMS) combined with tetranitro BT (TNBT) or nitro BT (NBT) or BSPT instead of ferri-ferrocyanide (tetrazolium method; McGADEY, 1967;LOJDA, 1976;GOSSRAU, 1978 c which can lead to false-negative results has to be kept in mind.…”

“…galactosidase hydrolyzes various natural (including lactose) and synthetic 8-galactosides but no 8-g1ucosides. ~10reover, the incubation medium is less stable than that used in the indigogenic method.With Fast Blue B instead of hexazotized new fuchsine or p-rosaniline the depiction of lysosomes is sometimes not so sharp.Acid B-galactosidase can also be demonstrated by means of a tetrazolium procedure(GOSSRAU, 1978 c). Kidney, liver, spleen, epididymis, uterus, and choroid plexus are especially rich in acid 8-galactosidase.…”

SERRS immunoassay for quantitative human CRP analysis

“…1. Bromochloroindolylphosphate (BCIP), a commonly used substrate for alkaline phosphatase, which generates a blue dimeric species with an absorption maximum of 615 nm in the presence of the enzyme, 25 was used as a substrate for an alkaline phosphatase (AP) conjugated anti-CRP antibody. Spectroscopic detection was accomplished using gold nanoparticles and excitation using 632.8 nm to create a SERRS spectrum with specific detection of a Raman shift at 600 cm -1 which was correlated to the concentration of CRP in standard samples following successful recognition of CRP.…”