Initial and Repetitive Yields from Proteins Blotted on PVDF Membranes

Abstract

PVDF (polyvinylidene difluoride) membranes have proven to be a useful support on which proteins or peptides can be immobilized during automated Edman degradation procedures (Matsudaira 1987). Several groups have reported that the initial and repetitive yields for samples electroblotted onto PVDF membranes (Baum et al 1987; Legendre and Matsudaira 1988; Moos et al 1988; Xu et al 1988; Yuen et al 1988) are much higher than samples blotted onto activated glass fiber filters (Vandekerchove et al 1985; Aebersold et al 1986). Since most of the measurements involved 100–500 pmole quantities of material, it would be valuable to evaluate the yields of < 100 pmole quantity of sample. In this study, I measured the initial and repetitive yields of 4–110 pmole quantities of 125I-s-lactoglobulin. The data suggests that both the repetitive and initial yields are independent of sample quantity and that sample loss during the sequencing cycles significantly affects the repetitive yields. Based on these results, it appears that sample detection is the major limit to routine sequence analysis of subpmole quantities of material.