Microfluidics as a high-throughput solution for chromatographic process development - The complexity of multimodal chromatography used as a proof of concept.

High-throughput technologies are fundamental to expedite the implementation of novel purification platforms. The possibility of performing process development within short periods of time while saving consumables and biological material are prime features for any high-throughput screening device. In this work, a microfluidic device is evaluated as high-throughput solution for a complete study of chromatographic operation conditions on ten different multimodal resins.

Correction: Enhancement of lateral flow assay performance by electromagnetic relocation of reporter particles.

[This corrects the article DOI: 10.1371/journal.pone.

Enhancement of lateral flow assay performance by electromagnetic relocation of reporter particles.

Lateral flow assays (LFAs) are a widely-used point-of care diagnostic format, but suffer from limited analytical sensitivity, especially when read by eye. It has recently been reported that LFA performance can be improved by using magnetic reporter particles and an external magnetic field applied at the test line. The mechanism of sensitivity/performance enhancement was suggested to be concentration/retardation of reporter particles at the test line.

High-Throughput Nanoliter-Scale Analysis and Optimization of Multimodal Chromatography for the Capture of Monoclonal Antibodies.

Multimodal ligands are synthetic molecules comprising multiple types of interactions that have been increasingly used for the capture of different biopharmaceutical compounds within complex biological mixtures. For monoclonal antibodies (mAbs) in particular, these ligands have shown the possibility of direct capture from cell culture supernatants in native conditions, as well as enhanced selectivity and affinity compared to traditional single-mode ligands. However, performing the capture of a target mAb using multimodal chromatography comes with the need for extensive optimization of the operating conditions, due to the multitude of interactions that can be promoted in parallel.

An overview of lectins purification strategies.

Lectins hold great promise not only as reagents for diagnostics and drug discovery but also as a novel class of biopharmaceutical products. In fact, new research directions in the last years have led to major developments in the uses of plant lectins as therapeutic agents against numerous diseases in an ageing society. It is even expected that lectins may occupy an important place in the biopharmaceutical industry next to monoclonal antibodies.

Formation of a misfolded conformation during refolding of HRPA1 in the presence of calcium.

Horseradish peroxidase A1 can refold to a native-like structure without binding calcium, originating a Ca2+-depleted native state as previously demonstrated. Thermal unfolding studies of horseradish peroxidase anionic 1 (HRPA1) have shown that calcium ions present during refolding lead to the appearance of a misfolded conformational state, which cannot incorporate the heme group. This calcium-induced conformational state, ICa2+, is less stable than the native state and has distinct secondary and tertiary structures as probed by far-UV and visible circular dichroism and tryptophan fluorescence.

Conformational states of HRPA1 induced by thermal unfolding: effect of low molecular weight solutes.

Fluorescence, CD, and activity measurements were used to characterize the different conformational states of horseradish peroxidase A1 induced by thermal unfolding. Picosecond time-resolved fluorescence studies showed a three-exponential decay dominated by a picosecond lifetime component resulting from energy transfer from tryptophan to heme. Upon thermal unfolding a decrease in the preexponential factor of the picosecond lifetime and an increase in the quantum yield were observed approaching the characteristics observed for apoHRPA1.

Heme and pH-dependent stability of an anionic horseradish peroxidase.

Horseradish peroxidase A1 thermal stability was studied by steady-state fluorescence, circular dichroism and differential scanning calorimetry at pH values of 4, 7 and 10. Changes in the intrinsic protein probes, tryptophan fluorescence, secondary structure, and heme group environment are not coincident. The T(m) values measured from the visible CD data are higher than those measured from Trp fluorescence and far-UV CD data at all pH values showing that the heme cavity is the last structural region to suffer significant conformational changes during thermal denaturation.

Enzymatic properties of a neutral endo-1,3(4)-beta-xylanase Xyl II from Bacillus subtilis.

A Bacillus sp. CCMI 966, characterised as Bacillus subtilis, has a duplication time of about 24 min. It produces at least two extracellular xylanases, Xyl I and Xyl II.