Validation of the drop coating deposition Raman method for protein analysis

Drop coating deposition Raman (DCDR) spectroscopy is critically evaluated to establish the limits to which it may be used to detect changes in protein conformation, binding, and purity. Difference spectroscopy is used to evaluate the reproducibility of the DCDR spectra under various experimental conditions. The results indicate (i) the absence of thermal/photochemical laser damage induced by the Raman excitation laser under typical DCDR data collection conditions, (ii) the reproducibility of DCDR spectra from samples with different volumes or concentrations, (iii) the water content of DCDR protein deposits and associated spectral signatures, and (iv) the degree of similarity between solution Raman spectra and DCDR spectra.