Thymidine Incorporation Assay
There are numerous methods available to study cell proliferation. The most reliable assays measure proliferation by directly measuring DNA synthesis.
The thymidine incorporation assay, the most common assay, utilizes a strategy wherein a radioactive nucleoside, 3H-thymidine, is incorporated into new strands of chromosomal DNA during mitotic cell division. A scintillation beta-counter is used to measure the radioactivity in DNA recovered from the cells in order to determine the extent of cell division that has occurred in response to a test agent.
More recently developed assays have eliminated the need to use radioisotopes. We offer many of these assays that provide significant advantages to the researcher.¹ Some of these products include 5-bromo-2'-deoxyuridine (BrdU), a nucleoside analog that is detected during active DNA synthesis using immunohistochemistry. We have also developed the Click-iT® EdU Microplate Assay that employs the nucleoside analog EdU (5-ethynyl-2’-deoxyuridine) and a detection method that is not antibody-based, so it does not require DNA denaturation.
The advantage of these incorporation assays is that they are direct measures of proliferation. Indirect methods such as those that measure mitochondrial activity (e.g. MTT) require additional confirmation since test agents could affect MTT processing without affecting cell viability. The user should be aware that identification of test agents that inhibit proliferation is not necessarily evidence of anti-angiogenic effect. In some cases, these agents may simply be toxic to the cell.
- Haugland, R. (2005). Assays for Cell Viability, Proliferation, and Function. In: The Handbook: A Guide to Fluorescent Probes and Labeling Technologies, 10th ed. USA: Invitrogen Corp. pp. 699-776.
LT170 updated 6-Oct-2011