Specialty Genotoxicity and Mutagenicity Service Analytics
Treat and Wash: Ames Test for Antibiotics, Antibodies, Proteins, Aminoacids and other compounds interfering in the regular Ames Test.
In the treat and wash method the test compound or control solution, bacteria and S9 mix for metabolic activation are mixed together for 90 min at 37°C. Wash solution is added and the washed bacteria are collected by centrifugation. Supernatant is discarded and the bacteria are resuspended in the residual solution prior to continue with the assay procedure for plate incorporation test.
In line with OECD 471, the Ames Mutagenicity Test is run on 5 different S.typhimurium strains - TA100, TA98, TA1535, TA1537, TA102 or on 4 S.typhimurium strains plus an E.coli strain - E.coli uvrA, E.coli pKM101 or E.coli uvrA(pKM101). If required and demanded, a prescreen assay is run to detect cytotoxic effect of the test compound in order to adjust dose range or solubility. The Ames test is run on agar plates using the Plate Incorporation Technology or Preincubatien Method, in presence and absence of metabolic activation.
Confirmation of the results can be obtained by a separate technology available from Xenometrix.
Service Analytics with different Ames Test Technologies :
- Miniaturized Ames Test : Screeninig of test compounds with Ames MPFTMminiaturized Assay with 10 mg test compound per strain or for the detection of genotoxic impurities, controlling concentrated water samples
- Ames Test on agar plate under GLP in compliance with OECD 471, REACH
- Ames MPF Aqua for testing environmental samples such as surface or waste water, soil, air, packing material in the food industry.
The synthesis of a pharmaceutically active ingredient involves the use of solvents, catalysts or other processing reagents. During chemical synthesis partial degradation of the compound or side products may arise. These impurities may be present in many pharmaceutical preparations. According to ICH guideline M7 genotoxic pharmaceuticals have to be tested for absence of genotoxic impurities.
Batch testing service of pharmaceuticals is offered with the traditional Ames agar plate test or with the validated, miniaturized Ames MPF, Ames II test systems, requiring only small amounts of test compound.
The γH2AX/pH3 in vitro genotoxicity assay is an optimal tool in the early discovery phase of drug development to assess genotoxic potential of chemicals, cosmetics, pharmaceuticals, agrochemicals, water samples ..etc. using litte amount of test compound and providing multiple information out the same experiment.-
The γH2AX/pH3 determines directly the genotoxic mode of action of test compounds by differentiation between aneugenic and clastogenic compounds using the In-Cell Western technology (ICW) in human cell lines. Cytotoxicity is detected simultaneously.
γH2AX assay is based on the phosphorylation of histone H2AX, a global indicator of genotoxicity. The assay allows to determine compounds with different mode of genotoxic action, such as DNA adducts, oxidative stress, alkylating agents, aneugenic agents, topoisomerase inhibitors, modification of the dNTP pool...
The Assay differenciates clearly between aneugenic (modifications in the number of chromosomes) and clastogenic (inducing DNA breaks) compounds. Aneugenic compounds induce either an increase or a decrease in pH3 depending on the mode of action. Clastogens induce γH2AX and cytotoxic compounds generate a decrease in these both biomarkers. Moreover, the use of different cell lines permits to discriminate directly from bioactivated genotoxins without the need of an exogenous metabolic activation system.
- Detection of phosphorylated histone ɣH2AX as global indicator of genotoxicity
- Detection of phosphorylation of histoneH3, a biomarker of aneugenic compounds.
- Identification of mode of action: aneugenic versus direct clastogenic or bioactivated clastogenic compound
- Simultaneous measurement of cytotoxicity
- Higher sensitivity as compared to Micronucleus Assay
- Can be used on a wide range of human cell lines,
- Screening Assay with a few mgs in a 96well plate format
The micronucleus test (MNT) is an in vitro tests for the assessment of genotoxicity of a chemical, pharmaceutical or an environmental test compound. By evaluating the presence of micronuclei the micronucleus test (MNT) identifies genotoxic compounds. Micronuclei may contain chromosome fragments produced from DNA breakage (clastogens) or whole chromosomes produced by disruption of the mitotic apparatus (aneugens). The resulting small micronuclei can be detected in the cytoplasm of interphase cells.
In vitro Micronucleus Test Service is run according to OECD 487 on TK6 cells or on human blood cells. DAPI staining and as a supplemental service telomere, centromere staining is performed to eliminate results in the grey zone.
It has been shown that telomere shortening can be in response to environemental and to chemical exposure and is associated with many health conditions. Recent publications show that exposure of mammalian cells to a toxic chemical is associated with shorthening telomere length (R M'kacher in press). The combination of the Micronucleus Assay and Telomere & Centromere staining offers an innovative technology to analyze chromosomal structure and provides multiple information of your test article in preclinical and clinical safety evaluation studies:
- Telomere and Centromere staining allows simultaneous and immediate discrimination between Clastogens and Aneugens
- Clear Results - No grey zone
- Micronucleus Assay can be run with TK6 cells or with blood lymhocytes according to OECD 487 under GLP
- Translation of the Assay system in other cell models
- Telomere and Centromere staining can also be performed in combination with the Chromosomal Aberration Test OECD 473
- An experienced team assists you in analysis and interpretation of results
The chromosome aberration test (CAT) evaluates the potential of a test compound to induce structural chromosomal abnormalities such as breaks and exchanges. The CAT is performed in primary human peripheral blood lymphocytes according to OECD 473 with DAPI staining or mFISH. Telomere and Centromere staining is offered as a supplemental service.
A positive outcome is characterized by a statistically significant, dose-dependent increase in aberrant cells that exceeds historical negative control limits.