Project description:Sample preparation is a crucial step in bottom-up proteomics. Analytical performances of bottom-up proteomics can be improved by the miniaturization of sample preparation steps. Many microfluidic devices are proposed in the field of proteomics. But many of them are not capable of handling complex sample and do not integrate the processing and digestion steps. We propose a ChipFilter Proteolysis (CFP) microfluidic device derived from the Filter Aided Sample Preparation FASP method for the miniaturization of protein processing and digestion steps in bottom-up proteomics. The microchip has two reaction chambers of 0.6 µL volume separated by a protein filtration membrane in regenerated cellulose. Cell lysis, protein concentration and rapid chemical and enzymatic treatment can be performed in our microfluidic device. Complex proteomic samples like yeast protein extract have already been analyzed with our microchip. Compared to the traditional FASP method, our microfluidic device offers a better proteome coverage with ten times less starting material and eight times quicker protocol.
Project description:Sample preparation is a crucial step in bottom-up proteomics. Analytical performances of bottom-up proteomics can be improved by the miniaturization of sample preparation steps. Many microfluidic devices are proposed in the field of proteomics. But many of them are not capable of handling complex sample and do not integrate the processing and digestion steps. We propose a ChipFilter Proteolysis (CFP) microfluidic device derived from the Filter Aided Sample Preparation FASP method for the miniaturization of protein processing and digestion steps in bottom-up proteomics. The microchip has two reaction chambers of 0.6 µL volume separated by a protein filtration membrane in regenerated cellulose. Cell lysis, protein concentration and rapid chemical and enzymatic treatment can be performed in our microfluidic device. Complex proteomic samples like yeast protein extract have already been analyzed with our microchip. Compared to the traditional FASP method, our microfluidic device offers a better proteome coverage with ten times less starting material and eight times quicker protocol.
Project description:Application of SPOT to Zn-treated HL-60 cells for multi-omics mechanism discovery. A time-resolved, multi-omics experiment using an aumtomated, unified sample preparation platform
Project description:Microfluidic deterministic barcoding of mRNAs and proteins in tissue slides followed by high throughput sequencing enables the construction of high-spatial-resolution multi-omics atlas at the genome scale. Applying it to mouse embryo tissues revealed major tissue (sub)types in early-stage organogenesis, brain micro-vasculatures, and the fine structure of an optical vesicle at the single-cell-layer resolution.
Project description:Omics research targets biomolecules such as proteins, lipids, and metabolites, providing a comprehensive view of biological systems. The field has seen significant advancements with the development of mass spectrometry (MS). However, the accuracy of quantitative analyses is crucial due to the expansion of omics applications. Accurate comparisons depend on sample preparation and normalization methods. Some biological samples, like tissues and feces, present inherent variations that challenge accuracy. Normalization aims to reduce these variations through pre-acquisition methods that equalize biomolecule content and post-acquisition methods that adjust instrument signals. Most research has focused on single-omics data and post-acquisition normalization. However, multi-omics research, essential for understanding complex biological systems, has not thoroughly evaluated normalization methods. This study evaluates three pre-acquisition normalization methods using methanol-chloroform-water extraction to enhance multi-omics data analysis. Our multi-omics data shows that the data is significantly different depending on the normalization methods and our optimized normalization method showed that there were significant multi-omics profile changes even with young mice tissue samples. Based on these findings, we suggest sample normalization based on total protein amount to minimize the sample variation and get an accurate comparison for multi-omics.
Project description:Here, we show that a ChipFilter microfluidic device coupled to LC-MS/MS can successfully be used for identification of microbial proteins. Using cultures of E. coli, B. subtilis and S. cerevisiae, we have shown that it is possible to directly lyse the cells and digest the proteins in the ChipFilter to allow higher number of proteins and peptides identification than standard protocols, even at low cell density. The peptides produced are overall longer after ChipFilter digestion but show no change in their degree of hydrophobicity. Analysis of a more complex mixture of 17 species from the gut microbiome showed that the ChipFilter preparation was able to identify and estimate the amount of 16 of these species.