An efficient strategy for cell-based antibody library selection using an integrated vector system

BACKGROUND: Cell panning of phage-displayed antibody library is a powerful tool for the development of therapeutic and imaging agents since disease-related cell surface proteins in native complex conformation can be directly targeted. Here, we employed a strategy taking advantage of an integrated vector system which allows rapid conversion of scFv-displaying phage into scFv-Fc format for efficient cell-based scFv library selection on a tetraspanin protein, CD9. RESULTS: A mouse scFv library constructed by using a phagemid vector, pDR-D1 was subjected to cell panning against stable CD9 transfectant, and the scFv repertoire from the enriched phage pool was directly transferred to a mammalian cassette vector, pDR-OriP-Fc1. The resulting constructs enabled transient expression of enough amounts of scFv-Fcs in HEK293E cells, and flow cytometric screening of binders for CD9 transfectant could be performed simply by using the culture supernatants. All three clones selected from the screening showed correct CD9-specificity. They could immunoprecipitate CD9 molecules out of the transfectant cell lysate and correctly stain endogenous CD9 expression on cancer cell membrane. Furthermore, competition assay with a known anti-CD9 monoclonal antibody (mAb) suggested that the binding epitopes of some of them overlap with that of the mAb which resides within the large extracellular loop of CD9. CONCLUSIONS: This study demonstrates that scFv-Fc from mammalian transient expression can be chosen as a reliable format for rapid screening and validation in cell-based scFv library selection, and the strategy described here will be applicable to efficient discovery of antibodies to diverse cell-surface targets

Measurement of the Background Activities of a 100Mo-enriched powder sample for AMoRE crystal material using a single high purity germanium detector

The Advanced Molybdenum-based Rare process Experiment (AMoRE) searches for neutrino-less double-beta (0{\nu}\b{eta}\b{eta}) decay of 100Mo in enriched molybdate crystals. The AMoRE crystals must have low levels of radioactive contamination to achieve low background signals with energies near the Q-value of the 100Mo 0{\nu}\b{eta}\b{eta} decay. To produce low-activity crystals, radioactive contaminants in the raw materials used to form the crystals must be controlled and quantified. 100EnrMoO3 powder, which is enriched in the 100Mo isotope, is of particular interest as it is the source of 100Mo in the crystals. A high-purity germanium detector having 100% relative efficiency, named CC1, is being operated in the Yangyang underground laboratory. Using CC1, we collected a gamma spectrum from a 1.6-kg 100EnrMoO3 powder sample enriched to 96.4% in 100Mo. Activities were analyzed for the isotopes 228Ac, 228Th, 226Ra, and 40K. They are long-lived naturally occurring isotopes that can produce background signals in the region of interest for AMoRE. Activities of both 228Ac and 228Th were < 1.0 mBq/kg at 90% confidence level (C.L.). The activity of 226Ra was measured to be 5.1 \pm 0.4 (stat) \pm 2.2 (syst) mBq/kg. The 40K activity was found as < 16.4 mBq/kg at 90% C.L.Comment: 20 pages, 6 figures, 5 table

Time-resolved pathogenic gene expression analysis of the plant pathogen Xanthomonas oryzae pv. oryzae

Virulence of wild-type and mutant Xoo strains on rice. (DOCX 16 kb

Unleashing the full potential of Hsp90 inhibitors as cancer therapeutics through simultaneous inactivation of Hsp90, Grp94, and TRAP1

Cancer therapeutics: Extending a drug&apos;s reach A new drug that blocks heat shock proteins (HSPs), helper proteins that are co-opted by cancer cells to promote tumor growth, shows promise for cancer treatment. Several drugs have targeted HSPs, since cancer cells are known to hijack these helper proteins to shield themselves from destruction by the body. However, the drugs have had limited success. Hye-Kyung Park and Byoung Heon Kang at Ulsan National Institutes of Science and Technology in South Korea and coworkers noticed that the drugs were not absorbed into mitochondria, a key cellular compartment, and HSPs in this compartment were therefore not being blocked. They identified a new HSP inhibitor that can reach every cellular compartment and inhibit all HSPs. Testing in mice showed that this inhibitor effectively triggered death of tumor cells, and therefore shows promise for anti-cancer therapy. The Hsp90 family proteins Hsp90, Grp94, and TRAP1 are present in the cell cytoplasm, endoplasmic reticulum, and mitochondria, respectively; all play important roles in tumorigenesis by regulating protein homeostasis in response to stress. Thus, simultaneous inhibition of all Hsp90 paralogs is a reasonable strategy for cancer therapy. However, since the existing pan-Hsp90 inhibitor does not accumulate in mitochondria, the potential anticancer activity of pan-Hsp90 inhibition has not yet been fully examined in vivo. Analysis of The Cancer Genome Atlas database revealed that all Hsp90 paralogs were upregulated in prostate cancer. Inactivation of all Hsp90 paralogs induced mitochondrial dysfunction, increased cytosolic calcium, and activated calcineurin. Active calcineurin blocked prosurvival heat shock responses upon Hsp90 inhibition by preventing nuclear translocation of HSF1. The purine scaffold derivative DN401 inhibited all Hsp90 paralogs simultaneously and showed stronger anticancer activity than other Hsp90 inhibitors. Pan-Hsp90 inhibition increased cytotoxicity and suppressed mechanisms that protect cancer cells, suggesting that it is a feasible strategy for the development of potent anticancer drugs. The mitochondria-permeable drug DN401 is a newly identified in vivo pan-Hsp90 inhibitor with potent anticancer activity

Wireless thin film transistor based on micro magnetic induction coupling antenna

A wireless thin film transistor (TFT) structure in which a source/drain or a gate is connected directly to a micro antenna to receive or transmit signals or power can be an important building block, acting as an electrical switch, a rectifier or an amplifier, for various electronics as well as microelectronics, since it allows simple connection with other devices, unlike conventional wire connections. An amorphous indium gallium zinc oxide (α-IGZO) TFT with magnetic antenna structure was fabricated and studied for this purpose. To enhance the induction coupling efficiency while maintaining the same small antenna size, a magnetic core structure consisting of Ni and nanowires was formed under the antenna. With the micro-antenna connected to a source/drain or a gate of the TFT, working electrical signals were well controlled. The results demonstrated the device as an alternative solution to existing wire connections which cause a number of problems in various fields such as flexible/wearable devices, body implanted devices, micro/nano robots, and sensors for the &apos;internet of things&apos; (IoT).1

Measurement of the Background Activities of a 100Mo-enriched Powder Sample for an AMoRE Crystal Material by using Fourteen High-Purity Germanium Detectors

The Advanced Molybdenum-based Rare process Experiment in its second phase (AMoRE-II) will search for neutrinoless double-beta (0{\nu}\b{eta}\b{eta}) decay of 100Mo in 200 kg of molybdate crystals. To achieve the zero-background level in the energy range of the double-beta decay Q-value of 100Mo, the radioactive contamination levels in AMoRE crystals should be low. 100EnrMoO3 powder, which is enriched in the 100Mo isotope, is used to grow the AMoRE crystals. A shielded array of fourteen high-purity germanium detectors with 70% relative efficiency each was used for the measurement of background activities in a sample of 9.6-kg powder. The detector system named CAGe located at the Yangyang underground laboratory was designed for measuring low levels of radioactivity from natural radioisotopes or cosmogenic nuclides such as 228Ac, 228Th, 226Ra, 88Y, and 40K. The activities of 228Ac and 228Th in the powder sample were 0.88 \pm 0.12 mBq/kg and 0.669 \pm 0.087 mBq/kg, respectively. The activity of 226Ra was measured to be 1.50 \pm 0.23 mBq/kg. The activity of 88Y was 0.101 \pm 0.016 mBq/kg. The activity of 40K was found as 36.0 \pm 4.1 mBq/kg.Comment: 24 pages, 10 figures, 5 table

Depression and suicide risk prediction models using blood-derived multi-omics data

More than 300 million people worldwide experience depression; annually, ~800,000 people die by suicide. Unfortunately, conventional interview-based diagnosis is insufficient to accurately predict a psychiatric status. We developed machine learning models to predict depression and suicide risk using blood methylome and transcriptome data from 56 suicide attempters (SAs), 39 patients with major depressive disorder (MDD), and 87 healthy controls. Our random forest classifiers showed accuracies of 92.6% in distinguishing SAs from MDD patients, 87.3% in distinguishing MDD patients from controls, and 86.7% in distinguishing SAs from controls. We also developed regression models for predicting psychiatric scales with R2 values of 0.961 and 0.943 for Hamilton Rating Scale for Depression???17 and Scale for Suicide Ideation, respectively. Multi-omics data were used to construct psychiatric status prediction models for improved mental health treatment