In summary, a simple and convenient method for the determination of OD260 of oligonucleotides using HPLC peak area is described. Although only quantification of ODN is described here, it is conceivable that the method can be equally applicable to RNA quantification. Because synthetic oligonucleotides typically have to go through the HPLC procedure for the purposes of purification or analysis, the direct OD260 determination method can save time for researchers by bypassing the step of measuring UV absorbance using a spectrometer.