![]() Other approaches to screen combinatorial libraries comprise the use of phage display and similar biological methods. On this silicon plate, fluorescence screening and sequencing by matrix-assisted laser desorption/ionization (MALDI)-MS/MS were combined. A capacity of 800 beads per chip was accomplished. In a lithographic process, a microstructured microwell array was manufactured, entrapping a single bead in each silicon well. developed chip-based techniques to combine affinity experiments and the MS decoding of the peptide sequence. published a sophisticated confocal nanoscanning and bead picking method (CONA), followed by sequencing of the binding peptides separately on a µHPLC/mass spectrometry (MS) instrument. For the automated isolation of positive hits from large OBOC libraries, Hintersteiner et al. Conventional FACS systems may also be used for the same purpose, keeping in mind that these devices are designed for analyzing cells in the size up to 10 µm. Alternatively, the separation can be done by sorting beads using a system designated as complex object parametric analyzer and sorter (COPAS), a modified flow cytometer (FACS), which is compatible with beads typically used in OBOC libraries with a diameter in the range of 70–90 µm. ![]() A strategy for analyzing large libraries is bulk separation by protein-coated magnetic particles. To avoid these complex and laborious steps, several improvements of the OBOC technology have been presented. In the present format, more than 30,000 beads can be screened on one slide. The new technique was validated by employing a FLAG-based model system, identifying new peptide binders for the monoclonal M2 anti-FLAG antibody, and was finally utilized to search for IgG-binding peptides. The use of an optimized ladder sequencing approach improved the accuracy of the de-novo sequencing step to nearly 100%. In a fluorescence scan followed by matrix-assisted laser desorption/ionization (MALDI)-time of flight (TOF) mass spectrometry, high-affinity binders are directly and unambiguously sequenced with high accuracy without picking of the positive beads. ![]() Subsequently, the chip is incubated with a fluorophore-labeled target protein. ![]() On a traditional glass slide provided with an electrically conductive tape, beads of a combinatorial peptide library are aligned and immobilized by application of a precision sieve. Herein, we present a high-throughput “ all-on-one chip” system to avoid slow and technically complex bead picking steps. The traditional peptide screening involves tedious steps such as affinity selection, bead picking, sequencing, and characterization. The demand for binders with high affinity and specificity towards various targets has surged in the biomedical and pharmaceutical field in recent years. Screening of one-bead-one-compound (OBOC) libraries is a proven procedure for the identification of protein-binding ligands. ![]()
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