Abstract
Nucleotide modifications constitute marks in RNA and DNA that contribute to gene regulation, development and other cellular processes. The understanding of their intricate molecular roles has been hampered by the high number of different modifications, the lack of effective methods and tools for their detection and quantification as well as by their complex structure-function relationship. The recent development of RNA and DNA immunoprecipitation followed by high-throughput sequencing (RIP- and DIP-seq) initiated detailed transcriptome- and genome-wide studies. Both techniques depend on highly specific and sensitive antibodies to specifically enrich the targeted modified nucleotides without background or potential biases. Here, we review the challenges and developments when generating and validating antibodies targeting modified nucleotides. We discuss antibody-antigen interactions, different strategies of antigen generation and compare different binder formats suitable for state-of-the-art high resolution mapping and imaging technologies.