Abstract
Surface Enhanced Raman Spectroscopy (SERS) has demonstrated significant benefit in the identification of biological molecules. In this paper we have examined how to identify and differentiate the 150 kDa protein myeloperoxidase (MPO) from its corresponding antibody (Ab) and their immunocomplex through the use of SERS. The SERS signal of these biological molecules was enabled by 40 nm gold nanoparticles. The SERS spectra for both MPO and the Ab (an IgG molecule) demonstrated results consistent with previous published work on the Raman spectra of MPO and IgG antibodies. The immunocomplex SERS spectra showed peak shifts and intensity variations that could be attributed to conformational changes that occur during immunocomplex formation. Several key spectral areas have been identified which correspond to specific amino acids being shielded from undergoing resonance while new amino acid residues are made visible in the SERS spectrum of the immunocomplex and could be a result of conformational binding. These results indicate that SERS can be used to identify binding events and distinguish an immunocomplex from its individual components.
Highlights
IntroductionThe strong intensities of Amide I and III bands indicate that these immunoglobulins are predominately composed of β-sheets
All chemicals were purchased from Sigma Aldrich unless otherwise mentioned, Ab was purchased from ABD-Serotec and MPO was purchased from Lee
The SERS spectrum of MPO (Fig. 1b) exhibits strong similarities to the Raman signals reported in previous studies [3,23,26,30,31]
Summary
The strong intensities of Amide I and III bands indicate that these immunoglobulins are predominately composed of β-sheets They compared the Raman signal of the lyophilized forms of rabbit anti-ovalbumin Ab and the immunocomplex of rabbit anti-ovalbumin Ab with the corresponding antigen, ovalbumin. Dou et al [6] used near infrared (excitation wavelength of 1064 nm) SERS to detect the conformational binding of anti-mouse IgG (bound to GNPs of 29.7 nm diameter) to mouse IgG antigen They observed significant increase in the amide I and III bands at 1645 and 1261 cm−1, respectively (similar to the work of Painter et al [24]), when the immunocomplex was formed and an increase in tryptophan residues at 1467, 1112 and 880 cm−1. We have identified bands that are unique to the immunocomplex of MPO and its Ab and explore the use of such bands as signatures of binding events
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