A Strategy for Monoclonal Antibody Production to Ecto-ATPases

Abstract

A variety of ecto-enzyme activities exist on the surfaces of all cells. The ecto-nucleotidases, ecto-ATPase, ecto-ADPase, and apyrase or ATP diphosphohydrolase (ecto-ATPDase), are of particular interest at this workshop. These enzymes are not as well understood as most of the other ecto-enzymes. There have been few attempts to purify these enzymes relative to a large number of descriptive studies. This is due in large part to the fact that these integral membrane glycoproteins (purification of which is particularly difficult) are rarely present in high concentration, and affinity chromatography of these enzymes has not always been successful. This paper will present a general strategy for production of monoclonal antibodies to these enzymes exploiting a new hybridoma screening assay. It will also discuss the use of the resulting monoclonal antibodies for immunopurification of the enzymes. The strategy and methods described have been applied to the purification of the chicken oviduct¹ and chicken liver² ecto-ATP diphosphohydrolase and to the chicken gizzard ecto-ATPase³. This paper will emphasize the methods used for purification of the chicken oviduct enzyme. Specific details for each enzyme have been published in the above references. This review will emphasize general methods which may be of use to other workers.

This paper is dedicated to the memory of Professor Rosenberg.

Deceased.