Disulfide linked dimers of apolipoprotein D in urine

Apolipoprotein (apo) D is a glycoprotein that contains at least one free cysteine. This allows the formation of disulfide linked dimers of apoD, a phenomenon that could interfere with the study of the isoforms of apoD. Consequently, it is important to consider the effects of hetero- and homodimer formation on the molecular heterogeneity of apoD as well as on the evaluation of the specificity of antibodies to this glycoprotein. The identification of apoD in urine has provided a potential new marker of tubular proteinuria. Thus, we have studied the specificity of our polyclonal antibodies to apoD against the proteins present in normal urine, and at the same time, the existence of dimeric species of apoD linked by disulfide bonds in urine. The specimens were obtained from apparently healthy individuals and analyzed by Western blot. The results showed that apoD in urine exists as a mixture of monomers and dimers, the latter having apparent molecular weights different from those occurring in plasma. Only monomeric apoD was observed under reducing conditions, proving the monospecificity of the polyclonal apoD antibodies.