BACKGROUND: Chromatin immunoprecipitation on tiling arrays (ChIP-chip) has been 
widely used to investigate the DNA binding sites for a variety of proteins on a 
genome-wide scale. However, several issues in the processing and analysis of 
ChIP-chip data have not been resolved fully, including the effect of background 
(mock control) subtraction and normalization within and across arrays.
RESULTS: The binding profiles of Drosophila male-specific lethal (MSL) complex 
on a tiling array provide a unique opportunity for investigating these topics, 
as it is known to bind on the X chromosome but not on the autosomes. These large 
bound and control regions on the same array allow clear evaluation of analytical 
methods.We introduce a novel normalization scheme specifically designed for 
ChIP-chip data from dual-channel arrays and demonstrate that this step is 
critical for correcting systematic dye-bias that may exist in the data. 
Subtraction of the mock (non-specific antibody or no antibody) control data is 
generally needed to eliminate the bias, but appropriate normalization obviates 
the need for mock experiments and increases the correlation among replicates. 
The idea underlying the normalization can be used subsequently to estimate the 
background noise level in each array for normalization across arrays. We 
demonstrate the effectiveness of the methods with the MSL complex binding data 
and other publicly available data.
CONCLUSION: Proper normalization is essential for ChIP-chip experiments. The 
proposed normalization technique can correct systematic errors and compensate 
for the lack of mock control data, thus reducing the experimental cost and 
producing more accurate results.