Protein quantification in ecological studies: A literature review and empirical comparisons of standard methodologies

Abstract Protein quantification is a routine procedure in ecological studies despite the inherent limitations of well‐acknowledged protein determination methods which have been largely overlooked by ecologists. Thus, we want to bridge this knowledge gap, in hopes of improving the way ecologists quantify proteins and interpret findings. We surveyed the ecological literature to determine how and why ecologists quantify proteins. To determine whether different quantification methods produce comparable results across taxa, and between populations of a single species, we estimated the protein content of eight phylogenetically diverse taxa, and of desert isopods fed different diets, using various derived protocols of the 'crude protein', Bradford and bicinchoninic acid approach (BCA) methods. We found that ecologists use many protein quantification procedures, often without reporting the crucial information needed to evaluate and repeat their methods. Our empirical work demonstrated that the three quantification methods examined, and their derived protocols, resulted in highly divergent protein estimations that were inconsistent in rank across taxa, preventing conversion between methods. We also found that different quantification methods yielded different answers to whether isopod protein content is affected by diet. We conclude that commonly used quantification techniques yield distinct protein estimations with varying precision, and no single method is likely to be more accurate than another across taxa which may lead to inconsistent results across taxa and between conspecifics. Inaccurate protein quantification may explain the observed mismatch between organismal N and protein that has plagued some recent studies and that contradicts the principles of ecological stoichiometry. We recommend using a single BCA protocol to reduce inconsistencies across studies, until the promising amino acid analysis becomes more affordable, accurate and accessible to ecologists. Until then, ecologists should consider the abovementioned drawbacks of protein quantification methods and interpret their results accordingly.