PMC:4996395 / 870-4541 JSONTXT

{"target":"https://pubannotation.org/docs/sourcedb/PMC/sourceid/4996395","sourcedb":"PMC","sourceid":"4996395","source_url":"https://www.ncbi.nlm.nih.gov/pmc/4996395","text":"1. Introduction\nThe complexity of the human proteome requires high-throughput (HT) approaches to define its study. During the last decade, protein microarrays have emerged as a useful tool for the analysis of the proteome at large scale [1]. Currently, protein microarrays have been successfully applied in the study of biomarkers, post-translational modifications (PTMs) of proteins, and various types of interactions with proteins. In addition, they have shed light on the biological roles of proteins related to and involved in diseases [2,3,4].\nAlthough recent advances have improved the sensitivity and reproducibility of common and widespread proteomics technologies (such as 2D-GE, MALDI-TOF, or LC-MS/MS), they are not readily implemented in a HT format [5]. In contrast with other proteomic strategies, protein microarrays avoid the need for pre-fractionation of the sample. In fact, complex and non-fractionated proteome mixtures, such as serum, plasma, urine and tissue extracts, can be directly used for experimentation [2]. For this reason, protein microarrays offer a powerful technology for functional proteomics analysis in HT format.\nMicroarray technologies utilize densely printed micro- or nano-spots of capture ligands immobilized onto a solid support that are exposed to samples containing the corresponding binding molecules (often referred to as queries), allowing the simultaneous analysis of thousands of capture targets within the same assay [6,7]. Thus, protein array technology enables multiplex and highly sensitive protein assays capable of handling and resolving complex proteomes with limited available samples [5,7].\nIn general, protein microarrays can be prepared in different formats (planar, beads…) and wide diversity of content (from antibodies and recombinant proteins to cell lysates). In fact, researchers have classified protein arrays based on the format or the content; however, in practice, these differences are more related to nomenclature than methods.\nThe first critical step to build protein microarrays is to display proteins on a solid surface for the detection of their biochemical activities in a multiplex manner. Hence, this is considered one of the challenges in protein microarrays field because of the high variability in biochemical properties (such as oligomerization states, PTMs, stability, affinities and specificities, isoelectric point…).\nIn addition, protein production and purification in a HT manner with high yield can be challenging [8]. This is because cell-based expression systems and the protocols of purification to generate large quantities of proteins are usually very tedious, result in highly variable yields and do not guarantee the protein integrity. These issues represent one of the major bottlenecks in HT functional proteomics studies. In Nucleic Acids Programmable Protein Arrays (NAPPA), the proteins are synthetized from a DNA template directly onto the surface of the array and the nascent protein is captured at the same time by an affinity reagent [9] avoiding the vast majority of drawbacks mentioned above (Figure 1).\nFigure 1 Diagram of Nucleic Acid Programmable Protein Array (NAPPA). Bovine serum albumin is used for printing purified template DNA (including the protein of interest and a tag molecule) onto a slide together with an antibody that recognizes the specific tag. When the cell extract is added, the transcription and translation are initiated and the expressed protein is captured by the anti-tag antibody. Here, we briefly review NAPPA technology and its recent applications in the study of pathologies, discovery of biomarkers and also vaccine development.","divisions":[{"label":"Title","span":{"begin":0,"end":15}},{"label":"Figure caption","span":{"begin":3112,"end":3519}}],"tracks":[{"project":"2_test","denotations":[{"id":"27600221-12634796-69478156","span":{"begin":541,"end":542},"obj":"12634796"},{"id":"27600221-22438764-69478157","span":{"begin":543,"end":544},"obj":"22438764"},{"id":"27600221-25153794-69478158","span":{"begin":545,"end":546},"obj":"25153794"},{"id":"27600221-21803154-69478159","span":{"begin":763,"end":764},"obj":"21803154"},{"id":"27600221-12634796-69478160","span":{"begin":1033,"end":1034},"obj":"12634796"},{"id":"27600221-15701447-69478161","span":{"begin":1469,"end":1470},"obj":"15701447"},{"id":"27600221-21421460-69478162","span":{"begin":1471,"end":1472},"obj":"21421460"},{"id":"27600221-21803154-69478163","span":{"begin":1644,"end":1645},"obj":"21803154"},{"id":"27600221-21421460-69478164","span":{"begin":1646,"end":1647},"obj":"21421460"},{"id":"27600221-12948670-69478165","span":{"begin":2505,"end":2506},"obj":"12948670"},{"id":"27600221-18469824-69478166","span":{"begin":3041,"end":3042},"obj":"18469824"}],"attributes":[{"subj":"27600221-12634796-69478156","pred":"source","obj":"2_test"},{"subj":"27600221-22438764-69478157","pred":"source","obj":"2_test"},{"subj":"27600221-25153794-69478158","pred":"source","obj":"2_test"},{"subj":"27600221-21803154-69478159","pred":"source","obj":"2_test"},{"subj":"27600221-12634796-69478160","pred":"source","obj":"2_test"},{"subj":"27600221-15701447-69478161","pred":"source","obj":"2_test"},{"subj":"27600221-21421460-69478162","pred":"source","obj":"2_test"},{"subj":"27600221-21803154-69478163","pred":"source","obj":"2_test"},{"subj":"27600221-21421460-69478164","pred":"source","obj":"2_test"},{"subj":"27600221-12948670-69478165","pred":"source","obj":"2_test"},{"subj":"27600221-18469824-69478166","pred":"source","obj":"2_test"}]}],"config":{"attribute types":[{"pred":"source","value type":"selection","values":[{"id":"2_test","color":"#ec9993","default":true}]}]}}