Xtt Test Account Options
The XTT test is based on the cleavage of the yellow tetrazolium salt XTT to form an orange water soluble formazan product by dehydrogenase activity in the. Der XTT-Test ist ähnlich wie der MTT-Test ein Versuchsansatz, bei dem in in-vitro-Versuchen die Vitalität (in einigen Publikationen auch Viabilität genannt) von. Der XTT-Reduktionstest EZ4U ist für eine vergleichende Bewertung des zytotoxischen Effektes von Dentalwerkstoffen geeignet. Erforderlich für die Testung sind. Zellviabilität (Zelllebensfähigkeit, Lebendzellanteil, englisch cell viability) bezeichnet in der Diese Viabilität kann zum Beispiel im Neutralrot-Test durch die Aufnahme des Vitalfarbstoffes Neutralrot (engl. oder seinem Analogon XTT sowie die Luciferase-basierten Verfahren weisen über das Redoxpotential indirekt die. XTT-Test EZ4U. Für die Prüfung auf Zytotoxizität wurde der nichtradioaktive Testkit zur Zellproliferations- und. Zytotoxizitätsbestimmung EZ4U verwendet.
Der XTT-Reduktionstest EZ4U ist für eine vergleichende Bewertung des zytotoxischen Effektes von Dentalwerkstoffen geeignet. Erforderlich für die Testung sind. Zellviabilität (Zelllebensfähigkeit, Lebendzellanteil, englisch cell viability) bezeichnet in der Diese Viabilität kann zum Beispiel im Neutralrot-Test durch die Aufnahme des Vitalfarbstoffes Neutralrot (engl. oder seinem Analogon XTT sowie die Luciferase-basierten Verfahren weisen über das Redoxpotential indirekt die. Der hier vorgestellte XTT-Test dient der Bestimmung der metabolischen Aktivität Zellen und ist eine Weiterentwicklung des schon lange bekannten MTT-Tests.
The absorption of the colored formazan derivate of XTT converted by the microbes is a measure of the cellular vitality.
A high absorption value indicates high metabolic activity. Figure 2 Fig. Figure 3 Fig. Absorption was no longer measurable in concentrations from 3.
A reduced XTT conversion was observed in saliva biofilms which had been subjected to antimicrobial treatment with gaseous ozone and chlorhexidine.
There was no difference in the conversion of XTT using a 0. No difference could be seen in the scanning electron micrographs between the untreated biofilm and the biofilm treated with ozone.
In both samples, the cells appear plump and the biofilm has a loosely bound structure. The bacteria in the biofilm treated with chlorhexidine are damaged and the overall structure appears to be tighter Figure 5 Fig.
After treatment with ozone, parts of the biofilm were dyed red cells with damaged membranes. After CHX treatment, no green colored areas cells with intact membranes were identified.
Causing typical dental diseases, such as caries and periodontitis, biofilms complicate the elimination of microbes responsible for forming the biofilms by antimicrobial substances.
The objective of this study was to develop a biofilm model suitable for testing the efficacy of antimicrobial substances with non-culture-based detection of the vitality of saliva biofilms using XTT, and to prepare a suitable XTT assay.
Our study had several limitations. We used saliva of volunteers to create a practically relevant biofilm model. For better understanding of the interactions between bacteria and XTT, we performed our experiments on machined titanium discs to exclude material hydrophobicity, retention niches such as cavities and porosities into which the biofilm could adhere.
Titanium implants have been successfully used in dentistry and biofilms on titanium are the central problem in peri-implantitis.
Peri-implantitis of osseointegrated oral implants is not a monoinfection by single pathogens; rather, they show the characteristics of mixed infections.
Whereas the single species of a mixture of bacteria could not induce experimental abscesses, the combination of these species could do it [ 24 ].
Plaque biofilms containing multiple species of appropriate bacteria should be more relevant for studying dental diseases and antimicrobial efficacies [ 25 ].
The first step was the use of only aerobe cultivation methods. Most XTT assays are carried out aerobically. In the next step we will use a subgingival plaque biofilm under anaerobic conditions for XTT assays.
But studies also showed that the same microbiota that can be found around implants under anaerobic conditions also can be found around teeth under aerobic conditions [ 26 ], [ 27 ].
XTT is a colorless tetrazolium salt, which is converted into a colored, water-soluble formazan derivate by dehydrogenases, with succinate dehydrogenase being particularly important [ 23 ] as it plays a major role in the energy supply of each individual living cell [ 28 ].
Unlike other tetrazolium salts e. Since the color change in the solution can be directly determined by photometry, the XTT test permits a large number of test objects to be tested for their vitality very quickly.
What is new is the composition of the XTT staining solution applied. The saliva biofilm contains a large variety of different microbes Gram-positive, Gram-negative bacteria and fungi.
To convert XTT, these microbes require various additives which function as electron carriers. The standard additive for Candida spp.
For Gram-positive cocci bacteria, PMS is used in most instances [ 34 ], but sometimes menadione is also used [ 35 ].
In the case of Gram-negative rod-shaped bacteria, mainly menadione is applied [ 35 ], [ 36 ]. Our own preliminary unpublished investigations confirm these results on the suitability of menadione for Candida albicans and PMS for Streptococcus mutans and Streptococcus sanguinis as representatives in dentistry.
As for Pseudomonas aeruginosa , the combined application of menadione and PMS turned out to be suitable [ 37 ].
By means of a culture-based analysis of the saliva, Gram-positive cocci bacteria, i. McCluskey et al. It turned out, however, that a high cell count of at least 4.
The higher the number of metabolically active cells, the higher the colorimetric signal. In addition, the higher the metabolism of cells, the higher the signal.
Obviously, there is no linear relation between the number of cells and the colorimetric signal [ 38 ]. When other dyes FDA und Syto9 were used, the adsorption even remained constant [ 39 ].
After 48 h of biofilm formation, cell densities of ca. The amount of retained product may vary between planktonic bacteria and biofilms [ 38 ].
Moreover, biofilms are subjected to other conditions than are fresh bacteria suspensions. Many pathogens are persistent and, therefore, exhibit lower metabolic activity [ 40 ].
For this reason, a minimum number of pathogens cannot be determined from suspensions and biofilms in exactly the same manner.
In addition, planktonic cells can invest more energy in routine metabolism [ 30 ]. In our experiments, the absorption was 30 times higher after 48 h than after 24 h, i.
However, the different metabolic levels do not lead to a logarithmic increase of the XTT reduction. This XTT-related observation was also made by other research groups [ 39 ].
Reduced formazan formation was observed due to the antimicrobial treatment of the saliva biofilm.
Consequently, the XTT test is suitable for determining the efficacy of antimicrobial substances, especially for screening.
The low chlorhexidine concentration used has already been investigated by other researchers, who noted insufficient antimicrobial efficacy in the biofilms [ 41 ].
On the other hand, chlorhexidine proved to be slightly superior to ozone. This has also been published by other research groups who applied alternative methods [ 42 ].
However, in vivo Hauser-Gerspach et al. The scanning electron micrographs confirmed this result.
Following the ozone treatment, the morphology of the cells showed no differences. Although the biofilm treated with chlorhexidine appeared to be damaged compared to the control, only a few cells were morphologically deformed.
In spite of some disadvantages, XTT with the addition of menadione and PMS is a suitable method for determining the vitality in bacterial saliva biofilms and permits assessment of the efficacy of antimicrobial substances.
The assay is easy to perform, and allows a large number of test objects to be tested. It is particularly suited to screening various factors influencing the biofilm, such as antiseptics or other physical or chemical treatments, for instance, ozone, photodynamic therapy or atmospheric pressure plasma.
We thank Tina Dornquast and Hartmut Fischer for their excellent technical assistance. We also thank PD Dr. Lutz Netuschil for the critical discussions.
National Center for Biotechnology Information , U. GMS Krankenhhyg Interdiszip. Published online Apr 4.
Author information Copyright and License information Disclaimer. You are free to copy, distribute and transmit the work, provided the original author and source are credited.
This article has been cited by other articles in PMC. Abstract Objective: Many dental diseases are attributable to biofilms.
Keywords: biofilm model, saliva, S. Abstract Ziel: Viele Zahnerkrankungen sind auf Biofilme zurückzuführen. Introduction Bacterial infections play a specific role in dentistry.
Materials and method Cultivation of biofilms Biofilms were cultured on titanium discs 5 mm in diameter and 1 mm thick Straumann, Basel, Switzerland.
Antiseptic treatment with chlorhexidine Chlorhexidine digluconate was used as a 0. Open in a separate window. Figure 1.
Analysis For all experiments, at least eight test objects each were used. Results Cultivation of biofilms The cultivation procedure was constantly checked for cultures by determining the CFU and for reproducibility by microscopy.
Optimization of the staining solution The absorption of the colored formazan derivate of XTT converted by the microbes is a measure of the cellular vitality.
Figure 2. Determination of the measuring range Figure 3 Fig. Figure 3. Antimicrobial treatment A reduced XTT conversion was observed in saliva biofilms which had been subjected to antimicrobial treatment with gaseous ozone and chlorhexidine.
Figure 4. Figure 5. SEM micrograph: h mature saliva biofilm: A untreated, B after ozone treatment, C after chlorhexidine treatment.
Magnification 10, x. Figure 6. Magnification x. Discussion Causing typical dental diseases, such as caries and periodontitis, biofilms complicate the elimination of microbes responsible for forming the biofilms by antimicrobial substances.
Competing interests The authors declare that they have no conflict of interest. References 1. Bortolaia C, Sbordone L. I biofilm del cavo orale.
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Minerva Stomatol. Carlsson J. Bacterial metabolism in dental biofilms. Adv Dent Res. Biofilm susceptibility to antimicrobials.
Developmental and metabolic aspects of a monobacterial plaque of Streptococcus mutans C grown on human enamel slabs in an artificial mouth model.
Plaque Data. Caries Res. Enamel Data. Beighton D. The complex oral microflora of high-risk individuals and groups and its role in the caries process.
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Some of its limitations include that it does not account for total viability and it is not particularly sensitive to low-viability assays; however, it is known for its quick pace.
The "tadpoling" method can be used to measure culture viability accurately, which is what depicts its main separation from "frogging".
From Wikipedia, the free encyclopedia. Retrieved In Steinberg P ed. Journal of General Microbiology. Applied and Environmental Microbiology.
European Journal of Phycology. Stoddart MJ Mammalian Cell Viability: Methods and Protocols. Methods in Molecular Biology. New York, NY: Springer.The donors did not take any medication three months prior the study and did not have active carious lesions or periodontal disease. Results: The XTT method lends see more to testing the vitality of microorganisms in saliva biofilms. Cell proliferation was determined by BrdU proliferation assay. Second, one cannot make interstrain comparisons in the absence of detailed standardization, since different strains read more substrate with different capabilities. Antifungal susceptibility of Candida albicans biofilms on titanium discs with different surface roughness. Comparison of biofilms formed by Candida albicans and Candida parapsilosis on bioprosthetic surfaces. Colorimetric assays of cellular viability are important tools in the study of read article cell activity. After read article h, cells were stained with annexin V and survival cells were graphed. With the XTT test, cell proliferation and viability of the cells after treatment with the test item are determined colorimetrically. Da jede Methode Schwächen aufweist, werden teilweise fluoreszierende Doppelfärbungen zum gleichzeitigen Nachweis von Apoptose und Nekrose durchgeführt, wie z. Elution Test. Es ist eine Vielzahl von Tests zur Bestimmung der Zellviabilität auf dem Markt, die nach article source Messprinzipien arbeiten. Agar Diffusion Test. Dieser Farbumschlag kann in einem Spektralphotometer photometrisch gemessen und ausgewertet werden . Genotoxicity Testing. Kontakt Angebotsanfrage. Cytotoxicity Testing.