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PubMed1998

Cytotoxic action of 2-deoxy-D-glucose tetraacetate in tumoral pancreatic islet cells.

Tumoral insulin-producing cells of the RINm5F line were cultured for 8-96 h in the absence or presence of 2-deoxy-D-glucose (0.15-0.80 mM) or its tetraacetate ester (0.08-0.80 mM). Despite the fact that over a short incubation of 120 min the utilization of D-[5-3H]glucose and oxidation of D-[U-14C]glucose were not more markedly decreased by 2-deoxy-D-glucose tetraacetate than by the unesterified glucose analogue, the growth of the tumoral cells, as assessed by either the generation of formazan from 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide or direct cell counting, was more severely affected by the ester than by 2-deoxy-D-glucose. At a high concentration (0.80 mM), the ester even decreased the cell number below its initial value. No restoration of cell growth was observed when the cells were exposed for only 8 h to 2-deoxy-D-glucose tetraacetate (0.80 mM) and then further cultured for 64 h in the absence of the ester. These findings indicate that such an ester acts as a powerful cytostatic and cytotoxic agent in this tumoral cell line.

Adenoma, Islet Cell↗

PubMed1999

A new direct microscopy based method for evaluating in-situ bioremediation.

A new epifluorescent microscopy based method using 5-cyano-2, 3-ditolyl tetrazolium chloride (CTC) and 5-(4,6-dichlorotriazinyl) aminofluoroscein (DTAF) was developed for quantifying total microbial biomass and evaluating levels of microbial activity. CTC is a tetrazolium dye that forms fluorescent intracellular formazan when biologically reduced by components of the electron transport system and/or dehydrogenases of metabolically active bacteria. DTAF is a fluorescein-based fluorochrome that selectively stains bacterial cell walls thereby enabling quantification of total bacterial biomass. CTC can be used in conjunction with DTAF to provide the optical resolution necessary to differentiate metabolically active cells from inactive cells in microbial populations associated with subsurface soils. The CTC/DTAF staining method has been shown to be effective for quantifying the metabolic activity of not only aerobic bacteria, but also diverse groups of anaerobic bacteria. This method allows for the rapid quantification of total and active bacterial numbers in complex soil samples without enrichment or cell elution. In this study, CTC/DTAF staining was applied to evaluate in-situ microbial activity in petroleum hydrocarbon contaminated subsurface soils from Sites 3 and 13 at Alameda Point, CA. At each site, subsurface microbial activity at two locations within contaminated plumes were examined and compared to activity at two geologically similar but uncontaminated background locations. Significant bacterial populations were detected in all soils examined, and the biomass estimates were several orders of magnitude higher than those obtained by conventional culture-based techniques. Both the total bacterial concentrations and the numbers of active bacteria in soils from contaminated areas were substantially higher than those observed in soils from background locations. Additionally, the percentages of metabolically active bacteria in the contaminated areas were consistently higher than those detected in background areas, suggesting that the enhanced microbial activity was due to microbial contaminant degradation. Although conventional heterotrophic plate counts failed to show significant microbial activity at either of the sites, soil gas carbon dioxide and methane measurements confirmed that hydrocarbon contaminant degradation was occurring in both areas. The CTC/DTAF staining protocol proved to be a rapid, reliable, and inexpensive method to evaluate the progress of in-situ bioremediation.

Bacteria↗

PubMed1999

Diaphorase activity in sebaceous glands and related structures of the male red deer.

The localisation of diaphorase was visualised by light microscopy using the dye nitro blue tetrazolium and NADPH as substrates. Under appropriate conditions, diaphorase reduces this dye to a dark blue insoluble formazan. The enzyme was located at very low activity in many tissue and glandular structures of the deer, but at very much higher activity in sebaceous glands in the dermal velvet of the antler and skin, and in additional sebaceous gland-related structures in the ear canal, prepuce and tail (scent) gland. Within sebaceous glands, activity was greatest in the outermost layers of the acini, but decreased as the cells progressed and differentiated centripetally. There was little or no difference between the staining observed when NADH was used as a substrate, compared to NADPH. There was generalised staining (usually light) for glucose-6-phosphate dehydrogenase, lactate dehydrogenase, and glycerol-3-phosphate dehydrogenase. However, this staining was not specifically localised to sebaceous glands and related structures, showing that the observed activity in these structures was due to a diaphorase that was distinct from any of the dehydrogenase activities tested. The possible role of diaphorase in sebaceous development and secretion is discussed.

Adrenal Glands↗

PubMed2000

The effect of repeated amitriptyline and desipramine administration on cytokine release in C57BL/6 mice.

This study examines the effects of repeated amitriptyline and desipramine administration (10 mg/kg, IP) on the immunoreactivity of saline-injected C57BL/6 mice, as evaluated by the ability of splenocytes to reduce a tetrazolium salt to formazan (MTT test), to proliferate, and to produce cytokines, such as interleukin (IL)-1, IL-2, IL-4, IL-6, IL-10 and interferon gamma (IFN-gamma). Desipramine and amitriptyline administered for one or two weeks enhance the biochemical (estimated by MTT test) and proliferative activities of splenocytes. One and two weeks administration of desipramine significantly reduces the secretion of IL-4, an anti-inflammatory cytokine. Amitriptyline administration for four weeks stimulates the proliferative activity of splenocytes and enhances IL-2 bioactivity, whereas four weeks desipramine aministration does not change these parameters in comparison to saline treated control mice. Prolonged desipramine administration (seven and 28 days) significantly increased the bioactivity of IL-1. Four weeks of prolonged administration of amitriptyline and desipramine induces a significant increase in the secretion of IL-10, a cytokine with immunosuppressive and anti-inflammatory activities. The results show that the immunoregulatory effects of tricyclic antidepressants in C57BL/6 mice depend on the drugs used and on the duration of administration.

Amitriptyline↗

PubMed1997

AMPA neurotoxicity in cultured cerebellar granule neurons: mode of cell death.

Various forms of cell death induced by the glutamate receptor agonist, alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA), were analyzed by determining the capacity of cultured cerebellar granule cells to metabolize 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) into formazan, by measuring the leakage of lactate dehydrogenase (LDH), by using confocal microscopy to visualize propidium iodide staining of apoptotic nuclei, and by using field inversion gel electrophoresis (FIGE) for the detection of AMPA-produced cleavage of DNA into high molecular-weight fragments (50 kbp). All these measures indicated that stimulation of AMPA receptors may be involved in the neurotoxic effects of glutamate, and that AMPA-induced neurotoxicity in cerebellar granule cells display morphologically distinct features of both necrotic and apoptotic modes of cell death. In agreement with previous observations, a blockade of AMPA receptor desensitization was necessary to unmask AMPA-induced functional responses in cultured cerebellar granule neurons in vitro. Microfluorimetric measurements of free cytoplasmic calcium concentrations ([Ca2+]i) in single cerebellar neurons revealed that AMPA neurotoxicity was accompanied by a pronounced elevation of [Ca2+]i. Our current results add further evidence to the notion that glutamate-induced neurotoxicity in cerebellar granule cells is mediated not only through NMDA receptors but also through a direct activation of AMPA receptor-regulated cation channels.

Animals↗

PubMed2002

In situ detection of reactive oxygen species and nitric oxide production in normal and pathological tissues: improvement by differential interference contrast.

Reactive oxygen species (ROS), among which nitric oxide (NO) is currently included, play a plethora of (patho)physiological roles. Harman's free radical theory of aging put forth over 40 years ago received full support since then. A nitric oxide hypothesis of aging recently proposed by McCann, is very likely to be the object of widespread investigation in the near future. Therefore, the possibility of localizing at the (sub)cellular level under the light microscope the sites of ROS and NO production with simple and reliable methods appears as a powerful tool for analytic cytology and pathology. Various histochemical methods were developed for visualizing ROS production; a recently improved version to localize superoxide (and possibly also singlet oxygen), based on a DAB-Mn2+ -Co2+ reaction, appears very promising. Since the direct detection of NO is still very difficult, the action sites of NO are currently localized by the identification of NO synthase (NOS). The most widespread method to reveal the catalytic activity of NOS is that of demonstrating the fixation-resistant NADPH diaphorase activity with the tetrazolium salt method. We have improved this method by using a tetrazolium salt whose formazan particles are very thin and lipid insoluble (tetranitroblue tetrazolium, TNBT) and by including a tissue protectant, polyvinyl alcohol, in the incubation medium. Here significant examples of application of the DAB-Mn2+ -Co2+ technique for ROS and the TNBT-PVA method for NOS to normal liver and brain and to solid tumors are presented. We further document the usefulness of Nomarkis's differential interference contrast (DIC) to analyze wide tissue areas where ROS production or NOS activity is low or even nil. The improved version for NOS allowed for the first time to demonstrate NOS activity in liver fat-storing cells and in astrocyte-like cells in the brain.

Aging↗

PubMed1999

Multicenter evaluation of the MB-Redox medium compared with radiometric BACTEC system, mycobacteria growth indicator tube (MGIT), and Löwenstein-Jensen medium for detection and recovery of acid-fast bacilli.

MB-Redox is a new manual culture system designed for the recovery of mycobacteria from clinical specimens. It consists of a liquid medium (modified Kirchner medium) containing a redox indicator, a colorless tetrazolium salt, which is reduced to colored formazan by actively growing mycobacteria. Acid fast bacilli (AFB) are easily detected in the medium as pink to purple pinhead-sized particles. We report the results of a multicenter study (involving four Italian microbiology laboratories processing 2370 clinical specimens) aiming to evaluate the recovery rates of AFB and time required for their detection by using the MB-Redox medium. Two different protocols were set up: in Protocol A (1580 specimens) the performance of MB-Redox was compared with those of the radiometric BACTEC 460 TB system (B460) and Löwenstein-Jensen medium (L-J), whereas in Protocol B (790 specimens) it was compared with those of the Mycobacteria Growth Indicator Tube (MGIT) and L-J. A total of 213 mycobacteria were recovered, including 172 Mycobacterium tuberculosis complex (MTB) isolates and 41 nontuberculous mycobacteria (NTM) isolates. In Protocol A, recovery rates were 81% for MB-Redox system, 84% for B460 system, and 77% for L-J. In Protocol B the recovery rates by individual system were 87, 83, and 76% for MB-Redox, MGIT, and L-J, respectively. Differences in both the protocols were not statistically significant. The MB-Redox system plus L-J (Combination 1) recovered 94% of the isolates in Protocol A and 93% in Protocol B, while B460 plus L-J (Combination 2) and MGIT plus L-J (Combination 3) detected 91 and 89% of all mycobacteria isolates respectively. No statistically significant differences were found among the combinations. The mean time to detection of mycobacteria was 16.3 days in Protocol A and 19.1 days in Protocol B with the MB-Redox system, 22.4 and 25.9 days with L-J, 13.2 days with B460, and 18.2 days with MGIT. The contamination rates were 2.1, 2.0, 1.9, and 3.6 for MB-Redox, B460, MGIT, and L-J respectively. The MB-Redox is a reliable, nonradiometric system for growth and detection of mycobacteria. When used in combination with a solid medium it proved to be an effective replacement for B460. The MB Redox system is a labor-intensive method requiring much handling during the visual reading procedures.

Bacteriological Techniques↗

PubMed1986

Progressive loss of the macrophage respiratory burst in oxygen toxicity.

The respiratory burst of rat alveolar macrophages stimulated by a variety of agents declines as a function of time of exposure to hyperoxia. Previous studies have evaluated this effect in terms of the stimulated O(2) production of a population of cells. The present study was designed to determine whether this decline is due to a "turning off" of the respiratory burst activity of some cells within the alveolar macrophage population or a general suppression of the activity of all cells. The phorbol myristate acetate (PMA) initiated respiratory burst of individual rat alveolar macrophages was monitored using the reaction of nitroblue tetrazolium (NBT), which results in the formation of a precipitate on active cells. The formazan staining was evaluated using black and white photographs of the cells and comparison to a scale constructed from photographed cells of four differing intensities of staining. Frequency distributions indicated that when the respiratory burst capability in the population of alveolar macrophages is impaired approximately 50% by oxygen exposure and/or culture in plastic vessels with artificial media, there is a gradual shift in NBT reduction rather than an "all or nothing" mechanism, in which the distribution would have reflected a shift from darkly stained cells to the very lessened or negligible staining observed at the end stage of oxygen toxicity.

Animals↗

PubMed2003

In vitro cytotoxic and anti-inflammatory effects of myrrh oil on human gingival fibroblasts and epithelial cells.

Limited scientific studies suggest that myrrh (Commiphora molmol) has antibacterial and anti-inflammatory activities. This study determined myrrh oil (MO) cytotoxicity to human gingival fibroblasts and epithelial cells and its effect, measured by ELISA, on interleukin (IL)-1beta-stimulated IL-6 and IL-8 production. Cell viability and cytotoxicity were determined by metabolic reduction of a tetrazolium salt to a formazan dye (MTT assay) and by release of lactate dehydrogenase (LDH) from membrane damaged (LDH release assay) cells, respectively. Based on the MTT assay, 24- and 48-h exposures to </=0.001% MO had little effect on fibroblast and epithelial cell (24-h only) viability. At 48 h, 0.0005-0.001% MO decreased epithelial cell viability 30-50%. After 24 and 48 h, MO, at >/=0.005%, maximally decreased viability of all cell lines. In the LDH release assay, exposure to </=0.0001% MO caused <10% cytotoxicity to all cells. At 24 h, >/=0.0025% MO caused maximal cytotoxicity; </=0.001% MO caused 10-70% cytotoxicity. At longer exposure times, epithelial cells were more susceptible to cytotoxic effects of MO. There was little or no detectable IL-1beta-stimulated production of IL-6 or IL-8 by cells exposed to >/=0.0025% MO, probably reflective of loss of viability. At subtoxic MO levels (0.00001-0.001%), there was a significant reduction of IL-1beta-stimulated IL-6 and IL-8 production by fibroblasts, but not by epithelial cells.

Biological Assay↗

PubMed2002

Mitochondrial dysfunction is an early indicator of doxorubicin-induced apoptosis.

Generation of reactive oxygen species and mitochondrial dysfunction has been implicated in doxorubicin-induced cardiotoxicity. This study examined pro-apoptotic mitochondrial cell death signals in an H9C2 myocyte rat cell line and in isolated rat heart mitochondria exposed to doxorubicin. Mitochondrial and cellular viability were assessed using an MTT viability assay (formazan product formed by functional mitochondrial dehydrogenases) and calcein AM dye (fluoresces upon cleavage by cytosolic esterases). Mitochondrial dysfunction followed by cell death was observed using nM concentrations of doxorubicin. Significant doxorubicin-induced cell death was not apparent until after 6 h following doxorubicin exposure using the calcein AM assay. The involvement of apoptosis is evidenced by an increase in TUNEL (terminal (TdT)-mediated dUTP-biotin nick end labeling)-positive nuclei following doxorubicin treatment. Furthermore, doxorubicin administered to isolated mitochondria induced a rapid increase in superoxide production, which persisted for at least 1 h and was followed by increased cytochrome c efflux. In addition, caspase-3 activity was increased with doxorubicin administration in the H9C2 myocyte cell line. An oxidant-mediated threshold of mitochondrial death may be required for doxorubicin-induced apoptosis.

Animals↗

PubMed1992

Adherence to and cytotoxicity of Escherichia coli for eucaryotic cell lines quantified by MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide).

Adherence of Escherichia coli to human epithelial cells (HEp-2) was studied using MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) which is cleaved by enzymes of eucaryotic or procaryotic cells to formazan. This method allows to quantify adherence of Escherichia coli to HEp-2 cells and offers the advantage of assaying a large number of eucaryotic cells without using specific antisera or radioactive material. Furthermore, toxic effects of isolated hemolysin cloned in Escherichia coli onto a renal tubular cell line (LLC-PK1) was investigated by this method, showing reduced cellular viability of tubular cells after an incubation period of 10 to 20 min. MTT is therefore considered to be useful to assay the adherence of Escherichia coli to eucaryotic cells and to quantify toxic effects in eucaryotic cells induced by bacterial virulence factors.

Bacterial Adhesion↗

PubMed2001

Colorimetric determination of tocopheryl acetate (vitamin E) in pure form and in multi-vitamin capsules.

A simple, accurate and sensitive method for the determination of micro amounts of vitamin E (V(E)) in pure form and in multi-vitamin capsules is described. The method is based on the reduction of tetrazolium blue in slightly alkaline medium by vitamin E after extraction from aqueous EDTA medium with petroleum ether and transesterification. The oxidation reduction reaction occurs after 10 min of heating in a water bath at 90+/-2 degrees C, leading to the formation of a highly coloured formazan derivative. Different variables affecting the colour development were investigated and optimized. Absorbance measurements were made at 526 nm and the calibration graph was linear for 0.2-11.0 microg/ml of (V(E)) with relative precision of about 0.7-1.5% when the standard additions method is used.

Antioxidants↗

PubMed1997

Histopathological changes induced by zinc hydroxide in rat lungs.

Rat lungs were histologically examined at 1, 7, 14 and 28 days following a single intratracheal instillation of zinc hydroxide (1 mM). After one day of treatment, no confirmatory findings were noted. The zinc hydroxide injections were followed by an increase in proliferating cell nuclear antigen labeling indices in both alveolar macrophages and terminal bronchioles. After 7 days, the zinc hydroxide-treated lungs showed thickening of the interstitium with infiltration by alveolar macrophages, and an increase in the grade of Masson's trichrome staining (collagen fiber) in the alveolar interstitium. Thereafter, these morphological changes disappeared. The vehicle- and zinc sulfate (1 mM)-exposed lungs had no abnormalities at any time point. Formazan deposits in alveolar macrophages, formed as a result of nitro blue tetrazolium reduction, were increased in zinc hydroxide-treated lung slices, suggesting that zinc hydroxide stimulated super oxide anion generation from alveolar macrophages. These results show that zinc hydroxide can induce morphological alterations of rat lungs.

Animals↗

PubMed1998

In vitro toxicity of surfactants in U937 cells: cell membrane integrity and mitochondrial function.

Surface-active agents are components of many drugs and cosmetics. In order to examine their cytotoxic and membrane-toxic potential, various surfactants were examined in U937 cells using the tetrazolium reduction assay EZ4U, a modified XTT test, and the arachidonic acid release test (AART). EZ4U measures the ability of living cells to reduce a colorless tetrazolium salt to an orange water-soluble formazan derivative by mitochondrial dehydrogenases. [3H]arachidonic acid ([3H]AA) is rapidly incorporated into cell membrane phospholipids. Due to membrane disintegration or enzymatic catalysis, it is released into the cell culture medium and can be measured by scintillation technique. The results after 24-hour-exposure are as follow (CC50 microg/ml; RC50 microg/ml): benzalkonium chloride (0.6), Cremophor A25 (1.4; 5.9), sodium cetearyl sulfate (1.6; 19.9), Brij78 (1.6; 7.7), TEGO betaine E (3.0; 19.3), TEGO betaine CKD (4.1; 20.2), TEGO betaine L7 (7.5; 20.0), sodium dodecyl sulfate (7.5; 39.0), Triton X-100 (8.9; 110), polysorbate 80 (48.1; 491), soybean lecithin (7920; 19940).

Arachidonic Acid↗

PubMed2002

Anti-proliferative effect of 20-hydroxyecdysone in a lepidopteran cell line.

Ecdysteroids are steroid hormones involved in the epidermal growth of arthropods, controlling cell proliferation and further differentiation of target cells. The epidermal cell line IAL-PID2, established from imaginal discs of the Indian meal moth Plodia interpunctella kept its sensitivity to ecdysteroids in vitro, cells being able to respond to them by cytological and biochemical changes. When added to the culture medium, 20-hydroxyecdysone (20E) stopped cell proliferation and induced formation of epithelial-like aggregates. In order to better understand the cellular sequence of ecdysteroids signalling in epidermal cells we used the IAL-PID2 cell line for in vitro investigations of cytological events induced by the moulting hormone. After a 40 h serum deprivation, formazan assay (XTT) was routinely used to evaluate anti-proliferative effects of 20E during cell cycle. We established a more precise timing of the period of cell sensitivity to the hormone during the cell cycle, by the use of the mitotic index and the BrdU incorporation test. These in vitro assays were performed in parallel with the description of some hormone dependant cytological events, using immunofluorescent labelling with anti-beta tubulin/FITC antibodies and DNA staining.

Animals↗

PubMed1994

Effects of radiologic contrast media on human endothelial and kidney cell lines: intracellular pH and cytotoxicity.

RATIONALE AND OBJECTIVES: Radiologic contrast media (CM) are hyperosmotic compounds injected undiluted into a patient's blood, in which they contact endothelial cells. For some types of cultured cells, the application of a hyperosmotic stimulus may cause intracellular pH (pHi) acidification that is related to the extent of hyperosmolality and that ultimately influences cellular function. Accordingly, endothelial and kidney cells, two types of cells known to be exposed to CM effects, were treated at relevant iodine concentrations with various CM (320-1500 mOsm) to determine whether cell exposure to CM can disturb the pH(i) and to examine the contribution of CM to cellular cytotoxicity. METHODS: Ionic (n = 3) and nonionic (n = 3) CM were compared. Changes in the pH(i) of human vascular endothelial and kidney cell lines were monitored by use of a pH-sensitive fluorescent dye (2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein acetoxymethyl ester). The viability of cells treated with CM was determined by measuring the reduction of a tetrazolium salt (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenlytetrazolium bromide) to violet formazan, a reaction that requires the activity of mitochondrial dehydrogenase; this measurement was made with a microplate reader. RESULTS: The pH(i) of endothelial and kidney cells exposed to 40-60 mg of iodine per milliliter of CM showed acidification (approximately 0.2 pH unit). Within minutes, gradual pH(i) alkalinization to baseline values occurred. The return to baseline values was slower with ionic compounds than with nonionic CM (P < 0.001). Nonionic agents caused less cellular damage than did ionic CM. CONCLUSIONS: The pH(i) is involved in the immediate intracellular transduction of CM effects in vitro. The exposure of cells to ionic CM is more detrimental than is exposure to nonionic CM, as demonstrated by disturbances in the cytosolic pH and by long-term effects on cell viability.

Adenocarcinoma↗

PubMed2003

Activated Kupffer cells play an important role in intra-hepatic Th1-associated necro-inflammation in Concanavalin A-induced hepatic injury in mice.

BACKGROUND/AIMS: To examine whether or not activated Kupffer cells play an important role in intra-hepatic Th1-associated necro-inflammation in Concanavalin A (Con A)-induced hepatic injury in mice. METHODS: Con A was administered to Balb/c mice pretreated with or without gadolinium chloride (GdCl(3)). Kupffer cell activation was evaluated by their ability to produce superoxide anions in situ under liver perfusion with nitro blue tetrazolium (NBT). Hepatic concentration of cytokines was measured by ELISA and the mRNA expression of CXC chemokine receptor 3 (CXCR3) was evaluated by RT-PCR. Immunohistochemical detection of CD4 positive lymphocytes in the liver was also performed. RESULTS: GdCl(3)-pretreatment significantly (P<0.01) reduced the serum levels of alanine aminotransferase (ALT) in Con A-treated mice. Formazan deposition in Kupffer cells, the hepatic concentration of tumor necrosis factor-alpha and interferon-gamma, the mRNA expression of CXCR3 and the CD4 positive lymphocytes in the liver were decreased in GdCl(3)-pretreated mice as compared with those without GdCl(3)-pretreatment (P<0.05, respectively). CONCLUSIONS: Activated Kupffer cells, which produce superoxide anions, are involved in Con A-induced hepatic necro-inflammation in mice possibly through the activation of Th1-associated immune response mediated by CD4 and/or CXCR3 positive cells recruited into the liver.

Journal Article↗

PubMed2005

Tetrazolium dyes as tools in cell biology: new insights into their cellular reduction.

Tetrazolium salts have become some of the most widely used tools in cell biology for measuring the metabolic activity of cells ranging from mammalian to microbial origin. With mammalian cells, fractionation studies indicate that the reduced pyridine nucleotide cofactor, NADH, is responsible for most MTT reduction and this is supported by studies with whole cells. MTT reduction is associated not only with mitochondria, but also with the cytoplasm and with non-mitochondrial membranes including the endosome/lysosome compartment and the plasma membrane. The net positive charge on tetrazolium salts like MTT and NBT appears to be the predominant factor involved in their cellular uptake via the plasma membrane potential. However, second generation tetrazolium dyes that form water-soluble formazans and require an intermediate electron acceptor for reduction (XTT, WST-1 and to some extent, MTS), are characterised by a net negative charge and are therefore largely cell-impermeable. Considerable evidence indicates that their reduction occurs at the cell surface, or at the level of the plasma membrane via trans-plasma membrane electron transport. The implications of these new findings are discussed in terms of the use of tetrazolium dyes as indicators of cell metabolism and their applications in cell biology.

Animals↗