R5.6: Predictive Assays

Predictive or clonogenic assays are used to assess the response of cells to ionising radiation. They may be used to assess cell survival following radiation by measuring colony formation after a particular dose. Several types of assay exist. A control assay, where no radiation is given, must be used to determine the efficacy of the test and adjust the observed outcome. This is known (for in vitro colony assays) as plating effectiveness.

In Vitro Assay

If the tumour cells are able to grow on a glass plate (often with a growth medium of dead tumour cells or connective tissue) then the number of colonies that grow after a set period of time is a useful measure. Unfortunately, normal tissue fibroblasts in tumour samples often outgrow the tumour cells. This can be overcome by using a medium that inhibits the growth of anchorage requiring cells such as fibroblasts, and/or promotes the growth of tumour cells.
Short term in vitro assays are not as reliable due to heterogeneity of human tumour specimens, tumour cell clump formation, and the time taken for irradiated cells to die.

Spleen Colony Assays

If bone marrow cells are injected into a synergistic mouse recipient, colonies in the spleen will form from the surviving stem cells. The number of colonies can be counted to assess cell survival of bone marrow cells following irradiation. This technique is also used for some types of mouse lymphoma cells.

Lung Colony Assays

This is similar to a spleen colony assay. Transplanted mouse tumours will often grow more readily in the lung and the number of colonies that form on the lung surface is related to the number of surviving tumour cells.

Limiting-Dilution Assays

Different sizes of tumour colonies may be inoculated into the subcutaneous tissue of a creature (usually a mouse). The number and size of colonies which grow is then observed. By taking into account the size of the inoculated tumour colony, highly accurate readings of cell survival can be obtained (down to 1 x 10-6 surviving cells).

Non-Clonagenic Assays

These do not rely on growing tumour cells and measuring colony formation. They include measuring dividing cells by blocking cytokinesis, incorporation of radioactive cell components in dividing cells and by measuring DNA damage in cells directly.