Establishment of a Small-scale Western Blotting System Named as “Micro-Western Blotting” for Mammalian Ova Analysis

The purpose of the present study was to try to improve the western blotting method in order to reduce the number of sampling ova for protein detection. We established a small-scale western blotting system, named “micro-western blotting”, in which the width of each lane was 1 mm and 2 μl sample was applied to each lane in SDS-PAGE. In this method, only 4 porcine ova were required to detect and analyze the phosphorylation states of p34^cdc2, a catalytic subunit of maturation promoting factor, and two major components of the MAP kinase cascade, ERK and MEK. The number of ova required for protein detection in this method was about one tenth of that required for the normal-scale method and the resolution qualities were comparable with the normal-scale method. The present system might be useful for analyzing biological molecules in small and rare materials such as mammalian oocytes and embryos.