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Integrated drug discovery outsourcing

Oncology


Selvita has an extensive expertise in lead generation and optimization within the area of kinase inhibitors.

We provide selection and validation of in vitro cellular disease models, together with validation of target proteins in functional cellular assays. Additionally we provide production of custom made cancer cell lines for semi-high throughput screening.

Selvita's research team has expertise in provision of variety of established viability/proliferation assays, including:

MTS cell viability assay (multiplex format, dose response curves)
A main application is to assess the viability and the proliferation of cells. It can also be used to determine cytotoxicity of potential medicinal agents and toxic materials. In this assay the activity of enzymes that reduce MTS to formazan dyes is measured.

 

BrdU incorporation assay - proliferation assay (multiplex format, dose response curves)
5-bromo-2'-deoxyuridine (BrdU) is a synthetic nucleoside that is an analogue of thymidine which can be incorporated into the newly synthesized DNA of replicating cells. BrdU is commonly used in the detection of proliferating cells in tissue cultures.

FACS cell cycle analysis (asynchronous cell lines, synchronized cells in G1/S and M, polyploidy levels)
The distribution of the nuclear DNA content within a cell population can provide a great deal of information including effects of drugs and stimuli on cell cycle and polyploidy levels. Flow cytometry on synchronized cells is the method of choice when precise analysis of cell cycle in combination with the expression of antigen is needed. Mitotic index for evaluation of proliferation levels Mitotic index is defined as the ratio between the number of cells in mitosis and the total number of cells. Mitotic cells can be labeled by established protocols both in cell lines and tissues using antibodies against the nuclear antigen Ki67, H3(Ser10) and H3(Thr3).

As well as in series of established apoptosis/cell death assays, such as:

LDH (Lactate dehydrogenase activity) - cell death assay (multiplex format, dose response curves). Tissue and cells breakdown elevates levels of LDH, and therefore a measure of it indicates, e.g., hemolysis and cell death.

FACS early apoptosis detection assay (Annexin V/PI staining)
Dead cells, necrotic cells and cells in early stages of apoptosis can be detected and distinguished by binding of annexin V (a phosphatidylserine-recognizing protein) and analysis by flow cytometry.

PARP cleavage for detection of apoptosis
During apoptosis PARP protein is cleaved by caspase-3, and possibly other caspases. Established protocols enable detection of cleaved PARP in both cell culture and in vivo samples.

Caspase activity - apoptosis assay (labeled substrates)
Caspase activation is a cellular event associated with the induction of apoptotic death. Detection of caspase activity using fluorogenic caspase substrates provides a useful assay for analyzing one of the earliest known biochemical events associated with apoptosis.

Clonogenic survival assay
This technique measures ability of single cells to form colonies for studying the effectiveness of specific agents on the survival and proliferation of cells. It is frequently used in to determine the effect of drugs or radiation on proliferating tumor cells. Counting of cells is done using image analysis software. Senescence induction assay (in cells ß-Galactosidase activity assay) Cellular senescence is the phenomenon by which normal diploid cells lose the ability to divide. Senescence bypass is involved in the development of cancer and suggests that senescence may represent a tumor suppressor mechanism that could be exploited as a basis for cancer therapy. Increased ß-Galactosidase activity accompanies certain morphological changes in senescent cells and can be quantitatively measured in this assay.

Combination index for evaluation of synergy
Evaluation of drug-drug interaction is important in all areas of medicine. There is increasing interest in moving new drugs into clinical trials in potentially active combinations based on preclinical testing data. The combination index analysis provided quantitative estimation for evaluating drug interactions, which can be classified as synergistic (combinations demonstrating greater than the additive activity expected from each agent alone), additive, or antagonistic (drugs showing less activity in combination than expected from the sum of each agent alone). We also provide cell signaling profiling using Multiplex ELISA and Western-Blot analysis.

Oncology - in vivo assays