Characterization of eleven monoclonal antibodies (MAbs), raised to isolated sodium dodecyl sulfate (SDS)-treated Alzheimer's neurofibrillary tangles (ANT), has revealed the presence of at least two different epitopes. MAbs were tested for reactivity to ubiquitin and paired helical filaments (PHF) isolated by three different procedures. The effect of protease and/or alkaline phosphatase pretreatment on the reactivity of the MAbs with isolated PHF was also examined. All MAbs that had reacted strongly in the ELISA with sonicated SDS-treated ANT also immune decorated isolated PHF to varying degrees. Two MAbs exhibited a high reactivity to PHF: 3-39 and 5-25. MAb 3-39 was found to recognize a protease sensitive epitope. In contrast MAb 5-25 was found to consistently decorate isolated PHF in all preparations and exhibited a strong reactivity to ubiquitin, and the epitope in isolated PHF was not protease sensitive. Thus structural PHF after protease treatment and detergent treatment contain an antigenic site that is present in ubiquitin.