A monoclonal antibody recognizing human cancers with amplification/overexpression of the human epidermal growth factor receptor -- Data Supplement - Supporting Information -- Proceedings of the National Academy of Sciences

Supporting information for Jungbluth et al. (2003) Proc. Natl. Acad. Sci. USA, 10.1073/pnas.232686499

Fig. 6.
Western blot reactivity of purified 806 and DH8.3 with Triton X-100 cell extracts. 806 reacted with both wtEGFR (170 and 160 kDa) and ΔEGFR (140 kDa), whereas mAb DH8.3 reacted with ΔEGFR only. Lanes: 1, U87MG_ΔEGFR; 2, U87MG_wtEGFR; 3, A431. Cell lysates were resolved by SDS/PAGE 6% polyacrylamide Tris-glycine precast gels under reducing (5% β-mercaptoethanol) conditions, and antibodies were used at a concentration of 5 μg/ml. The reactivities with 200-, 130-, and 70-kDa proteins are nonspecific and due to the secondary reagents used. Specific binding was detected by biotinylated species-specific secondary Abs followed by streptavidin-conjugated alkaline phosphatase and visualized by using chemiluminescence. Two antibodies to wtEGFR, 528 and R1 (1, 2), did not stain in Western blot.

1. Waterfield, M. D., Mayes, E. L., Stroobant, P., Bennet, P. L., Young, S., Goodfellow, P. N., Banting, G. S. & Ozanne, B. (1982) J. Cell. Biochem. 20, 149–161.

2. Lin, C. R., Chen, W. S., Kruiger, W., Stolarsky, L. S., Weber, W., Evans, R. M., Verma, I. M., Gill, G. N. & Rosenfeld, M. G. (1984) Science 224, 843–848.