Discovery of a Unique Set of Dog-Seroreactive Proteins Using Nucleic Acid Programmable Protein Array.

Valley Fever (VF), caused by fungi in the genus , is a prevalent disease in southwestern and western parts of the United States that affects both humans and animals, such as dogs. Although the immune responses to infection with spp. are not fully characterized, antibody-detection assays are used in conjunction with clinical presentation and radiologic findings to aid in the diagnosis of VF.

Mucosal and systemic antigen-specific antibody responses correlate with protection against active tuberculosis in nonhuman primates.

Background: Increasing evidence supports that antibodies can protect against active tuberculosis (TB) but knowledge of potentially protective antigens, especially in the airways, is limited. The main objective of this study was to identify antigen-specific airway and systemic immunoglobulin isotype responses associated with the outcome of controlled latent Mycobacterium tuberculosis (Mtb) infection (LTBI) versus uncontrolled infection (TB) in nonhuman primates.

Methods: In a case-control design, using non-parametric group comparisons with false discovery rate adjustments, we assessed antibodies in 57 cynomolgus macaques which, following low-dose airway Mtb infection, developed either LTBI or TB.

Quantitative assessment of multiple pathogen exposure and immune dynamics at scale.

Serology reveals exposure to pathogens, as well as the state of autoimmune and other clinical conditions. It is used to evaluate individuals and their histories and as a public health tool to track epidemics. Employing a variety of formats, studies nearly always perform serology by testing response to only one or a few antigens.

Scaling eukaryotic cell-free protein synthesis achieved with the versatile and high-yielding tobacco BY-2 cell lysate.

Eukaryotic cell-free protein synthesis (CFPS) can accelerate expression and high-throughput analysis of complex proteins with functionally relevant post-translational modifications (PTMs). However, low yields and difficulties scaling such systems have prevented their widespread adoption in protein research and manufacturing. Here, we provide detailed demonstrations for the capabilities of a CFPS system derived from Nicotiana tabacum BY-2 cell culture (BY-2 lysate; BYL).

Comparative Analysis of Antimicrobial Antibodies between Mild and Severe COVID-19.

Patients with 2019 coronavirus disease (COVID-19) exhibit a broad spectrum of clinical presentations. A person's antimicrobial antibody profile, as partially shaped by past infection or vaccination, can reflect the immune system health that is critical to control and resolve the infection. We performed an explorative immunoproteomics study using microbial protein arrays displaying 318 full-length antigens from 77 viruses and 3 bacteria.

Identification of anti-Helicobacter pylori antibody signatures in gastric intestinal metaplasia.

Background: Chronic Helicobacter pylori infection may induce gastric intestinal metaplasia (IM). We compared anti-H. pylori antibody profiles between IM cases and non-atrophic gastritis (NAG) controls.

Serological profiling of Crohn's disease and ulcerative colitis patients reveals anti-microbial antibody signatures.

Background: The healthcare burden of inflammatory bowel disease (IBD) is rising globally and there are limited non-invasive biomarkers for accurate and early diagnosis.

Aim: To understand the important role that intestinal microbiota play in IBD pathogenesis and identify anti-microbial antibody signatures that benefit clinical management of IBD patients.

Methods: We performed serological profiling of 100 Crohn's disease (CD) patients, 100 ulcerative colitis (UC) patients and 100 healthy controls against 1173 bacterial and 397 viral proteins from 50 bacteria and 33 viruses on protein microarrays.

Comparative Microbiomics Analysis of Antimicrobial Antibody Response between Patients with Lung Cancer and Control Subjects with Benign Pulmonary Nodules.

Background: CT screening can detect lung cancer early but suffers a high false-positive rate. There is a need for molecular biomarkers that can distinguish malignant and benign indeterminate pulmonary nodules (IPN) detected by CT scan.

Methods: We profiled antibodies against 901 individual microbial antigens from 27 bacteria and 29 viruses in sera from 127 lung adenocarcinoma (ADC), 123 smoker controls (SMC), 170 benign nodule controls (BNC) individuals using protein microarrays to identify ADC and BNC specific antimicrobial antibodies.

Identification of Antibody Biomarker Using High-Density Nucleic Acid Programmable Protein Array.

A novel protein microarray technology, called high-density nucleic acid programmable protein array (HD-NAPPA), enables the serological screening of thousands of proteins at one time. HD-NAPPA extends the capabilities of NAPPA, which produces protein microarrays on a conventional glass microscope slide. By comparison, HD-NAPPA displays proteins in over 10,000 nanowells etched in a silicon slide.

Identification of anti-Epstein-Barr virus (EBV) antibody signature in EBV-associated gastric carcinoma.

Background: Around 10% of gastric carcinomas (GC) contain Epstein-Barr virus (EBV) DNA. We characterized the GC-specific antibody response to this common infection, which may provide a noninvasive method to detect EBV-positive GC and elucidate its contribution to carcinogenesis.

Methods: Plasma samples from EBV-positive (n = 28) and EBV-negative (n = 34) Latvian GC patients were immune-profiled against 85 EBV proteins on a multi-microbial Nucleic Acid Programmable Protein Array (EBV-NAPPA).

Immunoproteomic Profiles in Gastric Cancer.

Chronic infection is the major risk factor for gastric cancer (GC). However, only some infected individuals develop this neoplasia. Previous serology studies have been limited by investigating small numbers of candidate antigens.

Multi-platform Approach for Microbial Biomarker Identification Using as a Model.

The identification of microbial biomarkers is critical for the diagnosis of a disease early during infection. However, the identification of reliable biomarkers is often hampered by a low concentration of microbes or biomarkers within host fluids or tissues. We have outlined a multi-platform strategy to assess microbial biomarkers that can be consistently detected in host samples, using , the causative agent of Lyme disease, as an example.

Tandem Histone-Binding Domains Enhance the Activity of a Synthetic Chromatin Effector.

Fusion proteins that specifically interact with biochemical marks on chromosomes represent a new class of synthetic transcriptional regulators that decode cell state information rather than DNA sequences. In multicellular organisms, information relevant to cell state, tissue identity, and oncogenesis is often encoded as biochemical modifications of histones, which are bound to DNA in eukaryotic nuclei and regulate gene expression states. We have previously reported the development and validation of the "polycomb-based transcription factor" (PcTF), a fusion protein that recognizes histone modifications through a protein-protein interaction between its polycomb chromodomain (PCD) motif and trimethylated lysine 27 of histone H3 (H3K27me3) at genomic sites.

Multiplexed Nucleic Acid Programmable Protein Arrays.

Cell-free protein microarrays display naturally-folded proteins based on just-in-time synthesis, and have made important contributions to basic and translational research. However, the risk of spot-to-spot cross-talk from protein diffusion during expression has limited the feature density of these arrays. In this work, we developed the Multiplexed Nucleic Acid Programmable Protein Array (M-NAPPA), which significantly increases the number of displayed proteins by multiplexing as many as five different gene plasmids within a printed spot.

Identification of Antibody Targets for Tuberculosis Serology using High-Density Nucleic Acid Programmable Protein Arrays.

Better and more diverse biomarkers for the development of simple point-of-care tests for active tuberculosis (TB), a clinically heterogeneous disease, are urgently needed. We generated a proteomic () High-Density Nucleic Acid Programmable Protein Array (HD-NAPPA) that used a novel multiplexed strategy for expedited high-throughput screening for antibody responses to the proteome. We screened sera from HIV uninfected and coinfected TB patients and controls ( = 120) from the US and South Africa (SA) using the multiplex HD-NAPPA for discovery, followed by deconvolution and validation through single protein HD-NAPPA with biologically independent samples ( = 124).

Biophysical Characterization of a Vaccine Candidate against HIV-1: The Transmembrane and Membrane Proximal Domains of HIV-1 gp41 as a Maltose Binding Protein Fusion.

The membrane proximal region (MPR, residues 649-683) and transmembrane domain (TMD, residues 684-705) of the gp41 subunit of HIV-1's envelope protein are highly conserved and are important in viral mucosal transmission, virus attachment and membrane fusion with target cells. Several structures of the trimeric membrane proximal external region (residues 662-683) of MPR have been reported at the atomic level; however, the atomic structure of the TMD still remains unknown. To elucidate the structure of both MPR and TMD, we expressed the region spanning both domains, MPR-TM (residues 649-705), in Escherichia coli as a fusion protein with maltose binding protein (MBP).

SPRi determination of inter-peptide interaction by using 3D supramolecular co-assembly polyrotaxane film.

Accurate measurement of inter-peptide interactions is beneficial for in-depth understanding disease-related protein folding and peptide aggregation, and further for designing and selecting potential peptide drugs to the target antigen. Herein, we demonstrate a 3D polyrotaxane (PRX) surface for detecting peptides interactions by surface plasmon resonance imaging (SPRi). This surface is supramolecular self-assembly monolayer (SAM) structure fabricated by threading α-cyclodextrans (α-CD) through a linear polyethylene glycol (PEG) chain fixed on gold chip surface to form pseudopolyrotaxane, and further capping the pseudopolyrotaxane with bulky terminated group to form PRX film.

Recombinant expression, purification, and biophysical characterization of the transmembrane and membrane proximal domains of HIV-1 gp41.

The transmembrane subunit (gp41) of the envelope glycoprotein of HIV-1 associates noncovalently with the surface subunit (gp120) and together they play essential roles in viral mucosal transmission and infection of target cells. The membrane proximal region (MPR) of gp41 is highly conserved and contains epitopes of broadly neutralizing antibodies. The transmembrane (TM) domain of gp41 not only anchors the envelope glycoprotein complex in the viral membrane but also dynamically affects the interactions of the MPR with the membrane.

Exploration of panviral proteome: high-throughput cloning and functional implications in virus-host interactions.

Throughout the long history of virus-host co-evolution, viruses have developed delicate strategies to facilitate their invasion and replication of their genome, while silencing the host immune responses through various mechanisms. The systematic characterization of viral protein-host interactions would yield invaluable information in the understanding of viral invasion/evasion, diagnosis and therapeutic treatment of a viral infection, and mechanisms of host biology. With more than 2,000 viral genomes sequenced, only a small percent of them are well investigated.

Stable and sensitive silver surface plasmon resonance imaging sensor using trilayered metallic structures.

The silver surface plasmon resonance (SPR) sensor has long been explored due to its intrinsic sensitivity enhancement over the conventional single-layered gold SPR sensor. However, the silver SPR sensor has not been exploited for practical applications because of pronounced instability problems. We propose a novel gold-silver-gold trilayered SPR sensor chip, in which an extra buffer layer of gold is added between the silver and substrate adhesion layer (i.

Microfluidic assay without blocking for rapid HIV screening and confirmation.

The essential step for HIV spreading limitation is the screening tests. However, there are multiple disadvantages in current screening assays which need further confirmation test. Herein we developed a rapid HIV assay combining screening and confirmation test by using the microfluidic network assay.