The DNA Damage and Toxicology Core provides expertise, equipment and facilities to perform DNA damage and toxicology studies related to toxic or hypoxic tissue/cell injury in drug development, diseases or aging. In addition to offering standard cytotoxicity assays, the core can measure oxidative damage and quantify levels of apoptosis and necrosis in cells and tissues by using quantitative cytochemistry, immunocytochemistry techniques, and 3-D imaging.
- Toxicity testing in vitro and in vivo
- Isolation of DNA in damage-free conditions
- In vitro DNA fragmentation assays
- DNA fragmentation assays in cells (single-cell DNA fragmentation through ds- or ss-breaks by neutral or alkaline Comet assay) and tissues (in situ DNA fragmentation by TUNEL or ISEL assays) combined, if required, with quantitative immunocytochemistry and intracellular localization of cell death-associated proteins
- Assessment of direct oxidative DNA breaks and modifications
- Measurements of oxidative, membrane damage and apoptotic markers using quantitative immunocytochemistry
- Measurement of apoptotic DNA-degrading enzymes activities
The mechanisms of toxic tissue injury and cell death underlie numerous human diseases as well as the effects of pharmaceutical drugs and environmental toxins. These mechanisms are complex and vary with tissue structure, vascularization, spectrum of cell death proteins, nature of damaging agent, the strength, length and frequency of the exposure to the injury, and many other factors. Accordingly, cell death can take forms of apoptosis, necrosis, autophagy, mitotic catastrophe, as well as some intermediate or more specific forms of cell death including aponecrosis, oncosis, anoikis or abortosis. To approach this complexity, the best strategy for assessment of toxic tissue injury includes the use of: a) common methods that occur in all injuries independent of causes, mechanisms or the origin of tissue; b) methods that are quantitative; and c) approaches that allow, if necessary, combining observations with other methods to determine the mechanism of cell injury and death.
DNA fragmentation (i.e., accumulation of unprepared double-stranded DNA breaks) is a form of DNA damage, which indicates that cell death reached the point of no return and became irreversible. DNA fragmentation is a hallmark of all types of cell death regardless of the mechanism. To determine the mechanisms and molecules involved in cell death, DNA fragmentation assays can be easily combined with immunocytochemistry.
The Work Order and Interdepartmental Transfer Form contains a list of available services and prices.
Please read the following instructions to help us serve you better:
Scheduling: Services are prioritized by date of request.
Before starting: Please contact the core director before starting the experiment. We would like to agree up front on the procedures, controls and costs. Although the core cannot help with planning your entire experiment, we can make suggestions to ensure you obtain reliable results.
Histology samples: Core personnel are also available to advise you on how to prepare your samples for histology. We cannot be responsible for tissue blocks prepared by another lab. There are many technical problems that can arise from inappropriate treatment of tissues, which could affect the quality of staining. Therefore, we suggest that you let the core fix your tissues and prepare blocks. This will tremendously increase the likelihood of successful staining. Prior to staining, we need to know: a) the tissues or cells that you will use; b) how samples were treated before they were delivered to the core; and c) your experimental plan (i.e., what are controls and what are samples with maximal effect expected) to help us understand if the staining was done correctly and whether controls worked.
Image capturing: If the core performs image capturing, the investigator is advised to be present during the session. If you think that image analyses can be done by you or your associates, this is possible too.
Quantitative image analyses: The core’s expertise is in quantitative cytochemistry and immunocytochemistry. Although sometimes images are enough for a publication or proposal, the data look more assured if quantification with appropriate statistical analysis is performed. Quantification can be done by an automatic, semi-automatic or manual mode. If desired, the quantification can be done in the individual tissue or cellular compartments. Although quantification is time-consuming and costly, results often are not recognized without quantification. If quantitative image analysis is requested, the investigator will be provided with the raw data in a spreadsheet ready for statistical analysis. In addition, representative digital images will be provided.
Work order estimate: A work order estimate may be requested to inform you of the potential cost of the services ordered. Note that the cost can be shared between collaborating investigators or partially paid by another entity.
DNA Damage & Toxicology Core
Biomedical Research Building I, Room B324
4301 West Markham Street
Mail Slot 638
Little Rock, AR 72205
Alexei Basnakian, M.D., Ph.D., Director
Department of Pharmacology & Toxicology
UAMS College of Medicine