Immunoassays for the masses?

Researchers from the University of Chicago have developed a method for using self-assembled monolayers as a matrix for MALDI (matrix assisted laser desorption ionisation) mass spectrometry immunoassays. The new approach, dubbed self-assembled monolayers for matrix-assisted laser desorption ionisation time-of-flight mass spectrometry or SAMDI-TOF MS, is discussed in an early view article in Analytical Chemistry​​. The researchers write: "self-assembled monolayers that are designed for specific applications with MS offer a straightforward method for analysing protein variants and post-translational modifications in humoral fluids and hold much promise in phenotyping proteins in diseased samples or across patient populations."​ Several approaches exist for the label-free detection of biomolecules, most notably Surface Plasmon Resonance (SPR) and mass spectrometry. These devices are often known as immunosensors and work by immobilising a capture agent such as a specific antibody or antigen. SPR offers the benefits that it can measure low affinity protein interactions in real time and provides quantitative results as well as in situ​ kinetic measurements. While MS-based immunosensors are unsuitable for kinetic experiments, they do provide chemical information about the analyte as well as being able to discriminate between background species that may interact with the sensor. The researchers first coated glass microscope slides with titanium and gold before coating them with asymmetric disulphides that were functionalised to recognise either histadine-tagged protein A or protein G. These monolayers could be stored for several days at 4ºC prior to use although the monolayers had previously been shown to be stable for months at 37ºC. Antibodies for various antigens can then be bound to the protein A (or G) and the researchers managed to bind multiple antibodies to a single slide as a mixture. These included antibodies raised against the proteins cysC, subunits of haemoglobin, human serum albumin and transferrin. Human serum was then diluted using phosphate buffered saline (PBS) before spotting onto the surface-bound monolayers. MALDI-TOF MS analysis allowed detection of all of the antibodies from human serum and the researchers write that: "the application of SAMDI-TOF for the evaluation of multiple proteins offers a significant advantage for clinical diagnostics by significantly reducing the time and sample consumption per assay." ​ The technique was also applied to the detection of post-translational modifications of cysC that may act as biomarkers for multiple sclerosis. The technique showed that non-multiple sclerosis sufferers only had an unmodified form of cysC that weighs 13.3kDa while multiple sclerosis sufferers had a considerable amount ( >50 per cent) of a truncated form of cysC that weighs 12.5kDa.