We recommend the hybridization to run between 4 and 16 hours. However, we regularly run the hybridization overnight. For example, start the reaction at 4/5pm and then perform probe capture at 9 or 10am the following day. We have tested longer hybridization times only up to 24 hours. If you want to break in between, you can store the dried down library pool with COT-1 DNA and universal blockers overnight at 4˚C if you want to delay starting the hybridization reaction.