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Recent advances in the field of cancer biology have accelerated the discovery and development of novel biopharmaceuticals. At the forefront of these drug development efforts are high‐throughput screening, compressed timelines, and limited sample quantities, all characteristic of the discovery space. To meet program targets, large numbers of protein variants must be produced, screened, and characterized, presenting a daunting analytical challenge. Additionally, the higher‐order structure is paramount for protein function and must be monitored as a critical quality attribute. Matrix‐assisted laser desorption/ionization mass spectrometry has been utilized as an ultra‐fast, automatable, sample‐sparing analytical tool for biomolecules. Our group has published applications integrating hydrogen‐deuterium exchange mass spectrometry with matrix‐assisted laser desorption/ionization mass spectrometry for the rapid conformational characterization of small proteins, the current work expands this application to monoclonal and bi‐specific antibodies. This study demonstrates the ability of the methodology, matrix‐assisted laser desorption/ionization hydrogen‐deuterium exchange mass spectrometry, to detect conformational differences between bi‐specific antibodies from different expression hosts. These conformational differences were validated by orthogonal techniques including circular dichroism, nuclear magnetic resonance, and size‐exclusion chromatography hydrogen‐deuterium exchange mass spectrometry. This work demonstrates the utility of applying the developed methodology as a rapid conformational screening tool to triage samples for further analytical characterization. [ABSTRACT FROM AUTHOR]

At the forefront of chemistry and biology research, development timelines are fast‐paced and large quantities of pure targets are rarely available. Herein, we introduce a new framework, which is built upon an automated, online trapping‐enrichment multi‐dimensional liquid chromatography platform (TE‐Dt‐mDLC) that enables: 1) highly efficient separation of complex mixtures in a first dimension (1D‐UV); 2) automated peak trapping‐enrichment and buffer removal achieved through a sequence of H2O and D2O washes using an independent pump setup; and 3) a second dimension separation (2D‐UV‐MS) with fully deuterated mobile phases and fraction collection to minimize protic residues for immediate NMR analysis while bypassing tedious drying processes and minimizing analyte degradation. Diverse examples of target isolation and characterization from organic synthesis and natural product chemistry laboratories are illustrated, demonstrating recoveries above 90 % using as little as a few micrograms of material. [ABSTRACT FROM AUTHOR]

Palladium‐catalyzed Suzuki‐Miyaura (SM) coupling is widely utilized in the construction of carbon‐carbon bonds. In this study, nanoelectrospray ionization mass spectrometry (nanoESI‐MS) is applied to simultaneously monitor precatalysts, catalytic intermediates, reagents, and products of the SM cross‐coupling reaction of 3‐Br‐5‐Ph‐pyridine and phenylboronic acid. A set of Pd cluster ions related to the monoligated Pd (0) active catalyst is detected, and its deconvoluted isotopic distribution reveals contributions from two neutral molecules. One is assigned to the generally accepted Pd(0) active catalyst, seen in MS as the protonated molecule, while the other is tentatively assigned to an oxidized catalyst which was found to increase as the reaction proceeds. Oxidative stress testing of a synthetic model catalyst 1,5‐cyclooctadiene Pd XPhos (COD−Pd‐XPhos) performed using FeCl3 supported this assignment. The formation and conversion of the oxidative addition intermediate during the catalytic cycle was monitored to provide information on the progress of the transmetalation step. [ABSTRACT FROM AUTHOR]