Paired helical filaments (PHF) in Alzheimer disease are phosphorylated at multiple sites : ∗L.R. Murthy and K. Iqbal. Institute for Basic Research in Developmental Disabilities, Staten Island, NY 10314, USA

Abstract

Antisera were raised in rabbits to purified bovine tau and to isolated Alzheimer paired helical filaments (PHF) washed with sodium dodecyl sulfate (SDS). Both anti-tau and anti-PHF sera labeled at electron microscopic level PHF which had been isolated either by extraction with SDS or treatment with crude collagenase. On immunoblots all anti-tau and anti-PHF sera labeled bovine brain tau as well as the major 45- to 62-kDa PHF polypeptides which had been previously shown to co-migrate on SDS gels with normal human tau (J. Biol. Chem., 261 (1986) 6084–6089). All antisera labeled Alzheimer neurofibrillary tangles on tissue sections and the PHF polypeptides on immunoblots. Pretreatment with alkaline phosphatase had no effect on the immunostaining. The antisera did not react with ubiquitin, neurofilament triplet polypeptides and with the exception of one antiserum with tubulin and high-molecular weight microtubule-associated proteins. Absorption of tau antisera with tau and PHF and of PHF antisera with PHF resulted in complete removal of the tangles-staining antibodies. In case of the anti-PHF sera when absorbed with tau, only the staining of a certain tangles population, the dense type, was eliminated and that too at more than 20 times the amount needed for the anti-tau sera; the staining of the loosely packed type of tangles, presumably the final stage, gradually decreased but was not completely abolished. On immunoblots the tau-like major PHF bands remained labeled by the tau-absorbed anti-PHF sera. These studies indicate that tau in PHF is present in different states of modification and that these alterations in tau are in addition to the abnormal phosphorylation shown previously (Proc. Natl. Acad. Sci. U.S.A., 83 (1986) 4913–4917).