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The formation of disulfide bonds is critical for stabilizing protein structures and maintaining protein functions. It is important to understand the linkages between multiple cysteine residues within a protein. In this review, the analytical approaches using mass spectrometry (MS) for disulfide linkage assignment are classified and discussed. Enzymatic digestion under appropriate conditions followed by various MS detection strategies remains the primary method for cysteine linkage analysis. In-source decay (ISD) and electron transfer dissociation (ETD) have been used to generate significant peptide signals that indicate the identities of peptides involved in disulfide bonds. In addition, chemical labeling and software algorithms were also developed to facilitate the automation of disulfide bond analysis. For proteins with complex disulfide structure, methods involving partial reduction coupled with differential alkylation were demonstrated to be useful. In the past two decades, MS has become one of the most valuable tools for protein disulfide bond analysis. It provides irreplaceable information including the peptide backbone sequences as well as the cysteine connection pattern when coupling with appropriate sample preparations. The related approaches with their unique features can be applied for different aims such as structural characterization or functional studies of proteins.

Paper diagnostics are devices made of paper and cellulosic materials to recognize and quantify biomolecules and chemical agents affecting health. There has been a rapid increase in interest for paper-based diagnostic methods in recent years. This is because paper is cheap, widely available, easily engineered, biodegradable, combustible, biocompatible, sterilizable, hydrophilic, and easy to functionalize and process into diagnostic devices. Paper is a very attractive substrate to develop as a low-cost diagnostic platform. Paper can serve four functions in a diagnostic: 1) transport and measurement of sample and analytes, 2) reaction support, 3) separation of reactants from products, and 4) communication of results. When an aqueous analyte, such as blood, saliva, urine or feces is tested using a paper diagnostics analytical device (PAD), the fluid is driven by capillary flow induced by the porous and hydrophilic structure of the cellulose fibers, causing wicking through the paper inter-fiber space. Should hydrophobic barriers be formed onto or within paper, microfluidic systems can be created, allowing for the passive transport of the analyte. This article has four sections. The first presents paper as a composite and highlights its properties and attributes with respect to modern paper diagnostics needs. Functional printing is presented as a technology to manufacture paper diagnostics. The second section analyzes paper diagnostic design. PADs based on one dimension (1D), 2D and 3D flow, the methods of reporting, and the principles of detection are reviewed. The third section investigates applications in health and medicine for paper diagnostics in terms of clinical diagnostics, physiological disorders, and pathogenic diseases. The last section presents an analytical perspective of some of the critical issues. It is the objective of this article to analyze paper as a viable technology for producing low-cost medical analysis and to delimit the range of applications and potential best suited to paper diagnostics.