**4. Host cell reactivation (HCR) assay**

As mentioned above, DNA repair activity plays a critical role in maintaining genome integrity. Regardless the alterations of DNA repair genes at the levels of gene expression or DNA sequence, measurement of DNA repair activity can reflect the overall biological effects that are as consequences of these molecular changes and/or anticancer drug responses. Here we describe an easy and fast functional assay (HCR) to evaluate cellular DNA repair activity *in vivo*. This method uses a plasmid that can produce luciferase in mammalian cells as a reporter. We choose luciferase as a reporter since its characteristics of high sensitivity and wide dynamic linear range for quantification. Of course, other commonly used reporters, such as chloramphenicol acetyltransferase (CAT), secreted alkaline phosphatase (SEAP) or green fluorescent protein (GFP) can also be used.

The reporter is damaged *in vitro* first and is transfected into host cells. If the damaged reporter plasmid can be repaired in the host cells, the luciferase will be re-expressed. Otherwise, the luciferase activity will be much lower than that transfected with undamaged control plasmid. By this way, one can determine the DNA repair capacity by simply measuring luciferase activity. The reporter plasmid can be damaged using various methods such as UV, chemicals or restriction enzymes and serve as substrates for different DNA repair pathways. In this chapter, we will demonstrate the use of HCR in evaluating DNA repair capacities via NER, HR and NHEJ pathways.
