Surface plasmon resonance investigations of human epidermal growth factor receptor 2.
This investigation utilizes surface plasmon resonance (SPR) spectroscopy to detect and quantify human epidermal growth factor receptor 2 (HER-2), an oncogene product that is over-expressed in some aggressive forms of breast cancer. Specifically, the HER-2 trans-membrane protein p185 and its extra cellular fragment p105 are analytes targeted in this work by using a gold-based biosensor slide on which an anti-HER-2 antibody has been immobilized by attachment to Protein G that is fixed to the gold film. A detection limit of > or =11 ng/mL for p185 resulted when trastuzumab was used as the anti-HER-2 antibody on the biosensor slide. Experiments with semi-purified p105 revealed that it binds weakly and reversibly to trastuzumab, therefore complicating its detection and quantification. Results of studies that reacted a 13-amino-acid peptide (PP13) from the HER-2 kinase domain with its specific antibody were critically different than p185 and p105 studies. Spectral analysis of the reflectivity at constant bulk buffer refractive index revealed a progressive negative SPR shift over time. A negative shift suggests that a loss of protein mass from the anti-PP13 antibody-Protein G biosensor is occurring. Several possibilities that may explain these negative SPR shifts are discussed.
Martin
V S
VS
Department of Chemistry, University of Wyoming, Laramie, Wyoming 82071, USA.
Sullivan
B A
BA
Walker
K
K
Hawk
H
H
Sullivan
B P
BP
Noe
L J
LJ
D000911:Antibodies, Monoclonal / Q000737:chemistry; D061067:Antibodies, Monoclonal, Humanized; D015374:Biosensing Techniques; D045744:Cell Line, Tumor; D006046:Gold; D006801:Humans; D007118:Immunoassay; D009419:Nerve Tissue Proteins / Q000737:chemistry; D009842:Oligopeptides / Q000737:chemistry; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000032:analysis* / Q000276:immunology / Q000302:isolation & purification; D011994:Recombinant Proteins / Q000737:chemistry; D020349:Surface Plasmon Resonance; D000068878:Trastuzumab
D000911:Antibodies, Monoclonal; D061067:Antibodies, Monoclonal, Humanized; D006046:Gold; D009419:Nerve Tissue Proteins; D009842:Oligopeptides; D018719:Erb-b2 Receptor Tyrosine Kinases; D011994:Recombinant Proteins; D000068878:Trastuzumab; C029152:G-substrate