Publications

A multiplexed microarray chip (Immuno-mu SARS2) aiming at providing information on the prognosis of the COVID-19 has been developed. The diagnostic technology records information related to the profile of the immunological response of patients infected by the SARS-CoV-2 virus. The diagnostic technology delivers information on the avidity of the sera against 28 different peptide epitopes and 7 proteins printed on a 25 mm2 area of a glass slide. The peptide epitopes (12-15 mer) derived from structural proteins (Spike and Nucleocapsid) have been rationally designed, synthesized, and used to develop Immuno-mu SARS2 as a multiplexed and high-throughput fluorescent microarray platform. The analysis of 755 human serum samples (321 from PCR+ patients; 288 from PCR- patients; 115 from prepandemic individuals and classified as hospitalized, admitted to intensive-care unit (ICU), and exitus) from three independent cohorts has shown that the chips perform with a 98% specificity and 91% sensitivity identifying RT-PCR+ patients. Computational analysis utilized to correlate the immunological signatures of the samples analyzed indicate significant prediction rates against exitus conditions with 82% accuracy, ICU admissions with 80% accuracy, and 73% accuracy over hospitalization requirement compared to asymptomatic patients' fingerprints. The miniaturized microarray chip allows simultaneous determination of 96 samples (24 samples/slide) in 90 min and requires only 10 mu L of sera. The diagnostic approach presented for the first time here could have a great value in assisting clinicians in decision-making based on the information provided by the Immuno-mu SARS2 regarding progression of the disease and could be easily implemented in diagnostics of other infectious diseases.

JTD Keywords: Antibodies, Clinical diagnostic, Diagnosis, High-throughput, Machine learning, Microarray, Multiplexation, Nucleocapsid protein, Peptide epitopes, Sars-cov-, Sars-cov-2, Serological signature, Seroprevalence, Severity prediction, Spik

Plaques of the amyloid beta (A beta) peptide are a pathological hallmark of Alzheimer's disease (AD), the most common form of dementia. Mutations in A beta also cause familial forms of AD (fAD). Here, we use deep mutational scanning to quantify the effects of >14,000 mutations on the aggregation of A beta. The resulting genetic landscape reveals mechanistic insights into fibril nucleation, including the importance of charge and gatekeeper residues in the disordered region outside of the amyloid core in preventing nucleation. Strikingly, unlike computational predictors and previous measurements, the empirical nucleation scores accurately identify all known dominant fAD mutations in A beta, genetically validating that the mechanism of nucleation in a cell-based assay is likely to be very similar to the mechanism that causes the human disease. These results provide the first comprehensive atlas of how mutations alter the formation of any amyloid fibril and a resource for the interpretation of genetic variation in A beta.

JTD Keywords: aggregation, kinetics, oligomers, onset, rates, state, Aggregation, Alzheimer disease, Alzheimer's, Amyloid, Amyloid beta-peptides, Computational biology, Deep mutagenesis, Dna mutational analysis, Genetics, Genomics, High-throughput nucleotide sequencing, Kinetics, Mutation, Nucleation, Oligomers, Onset, Plasmids, Precursor protein, Rates, S. cerevisiae, Saccharomyces cerevisiae, State, Systems biology

Here we describe the design and evaluation of a fluidic device for the automatic processing of microarrays, called microarray processing station or MPS. The microarray processing station once installed on a commercial microarrayer allows automating the washing, and drying steps, which are often performed manually. The substrate where the assay occurs remains on place during the microarray printing, incubation and processing steps, therefore the addressing of nL volumes of the distinct immunoassay reagents such as capture and detection antibodies and samples can be performed on the same coordinate of the substrate with a perfect alignment without requiring any additional mechanical or optical re-alignment methods. This allows the performance of independent immunoassays in a single microarray spot.

JTD Keywords: Automation, Customization, High-throughput screening, Immunoassays, Microarrays

Xenopus tadpoles are an emerging model for developmental, genetic and behavioral studies. A small size, optical accessibility of most of their organs, together with a close genetic and structural relationship to humans make them a convenient experimental model. However, there is only a limited toolset available to measure behavior and organ function of these animals at medium or high-throughput. Herein, we describe an imaging-based platform to quantify body and autonomic movements of Xenopus tropicalis tadpoles of advanced developmental stages. Animals alternate periods of quiescence and locomotor movements and display buccal pumping for oxygen uptake from water and rhythmic cardiac movements. We imaged up to 24 animals in parallel and automatically tracked and quantified their movements by using image analysis software. Animal trajectories, moved distances, activity time, buccal pumping rates and heart beat rates were calculated and used to characterize the effects of test compounds. We evaluated the effects of propranolol and atropine, observing a dose-dependent bradycardia and tachycardia, respectively. This imaging and analysis platform is a simple, cost-effective high-throughput in vivo assay system for genetic, toxicological or pharmacological characterizations.

JTD Keywords: Xenopus tropicalis, Animal behavior, Cardiac imaging, Motion analysis, Animal tracking, Hhigh-throughput in vivo assay