Available Services

Zeiss LSM980-NLO confocal microscope with multiphoton excitation and Airyscan2 detector.

• Multiphoton excitation: Coherent Discovery NX (dual line, tunable 700-1300 nm and fixed 1040 nm)
• Available laser sources: 405 nm, 488 nm, 561 nm, 594 nm, 639 nm
• Non-descanned detectors
• Airyscan2 detector
• Incubator (CO₂, temperature)
• Epifluorescence (sCMOS camera Zeiss 702)
• Zen Blue version 9
• Available objectives:
• 5x/0.085 EC Plan-Neofluar
• 10x/0.45 Plan-Apochromat
• 20x/0.8 Plan-Apochromat DIC II
• 40x/1.4 Oil Plan-Apochromat
• 40x/1.1 Water C-ApoCHROMAT
• 63x/1.4 Oil PlanApo DIC II

Zeiss LSM980 confocal microscope with Airyscan2 detector.

• Available laser sources: 405 nm, 488 nm, 561 nm, 594 639 nm
• Airyscan2 detector
• Sample finder
• Incubator (CO₂, temperature)
• Epifluorescence (sCMOS camera Zeiss 702)
• Zen Blue version 9
• Available objectives:
  • 5x/0.12 Fluar
  • 10x/0.45 Plan-Apochromat
  • 20x/0.8 Plan-Apochromat
  • 40x/1.4 Oil Plan-Apochromat
  • 63x/1.4 Oil Plan-Apochromat

Zeiss Lattice Lightsheet 7 with dual cameras.

• Available lasers sources: 488 nm, 561 nm and 640 nm
• Dual cameras (Hamamatsu ORCA Fusion)
• Incubator (CO₂, temperature)
• Zeiss Zen Blue software (3.10)

Zeiss Elyra7 lattice structured illumination and localization microscopy system.

• Available laser sources: 405 nm, 488 nm, 561 nm, 638 nm
• Dual camera (PCO.EDGE)
• Incubator (CO₂, temperature)
• Dual computer with Zen black 0 SR for image acquisition and Zen Blue 3.7 for processing
• Available objectives:
• 10x/0.3 EcPlan DIC I
• 20x/0.8 PlanApo DIC II 40x/1.2 W Apo DIC III
• 63x/1.4 Oil PlanApo DIC I
• 63x/1.46 aPlanApo DIC III
• 100x/1.57 aPlanApo DIC III

The Zeiss Axioscan Z.1 is an automated slide scanner designed to capture whole slide images in both brightfield and epifluorescence modes, accommodating up to 100 slides per session.

Configuration:

• Cameras:
  • Hamamatsu Orca-Flash 4.0 v3 sCMOS mono camera
  • Hitachi HV-F203SCL 3CCD color camera
• Light sources:
  • Colibri 7 for epifluorescence (385 nm, 430 nm, 475 nm, 555 nm, 590 nm, 630 nm,

735 nm)

• White led lamp for transmitted light

• Filter sets optimised for:
  • DAPI (Zeiss BP 450/40)
  • GFP / Alexa Fluor 488 / FITC (Zeiss BP 525/50)
  • Cy3 / Alexa Fluor 555 / TRITC (Zeiss BP 605/70)
  • Cy5 / Alexa Fluor 647 / APC (Zeiss BP 690/50)
  • DAPI/GFP/Cy3/Cy5/Cy7 (Zeiss PBP 425/30+514/31+592/25+681/45+785/38)
• 6 objectives:
  • 2,5x / 12 NA Fluar M27
  • 5x / 25 NA Fluar M27
  • 10x / 45 NA Plan-Apochromat M27
  • 20x / 45 NA N-Achroplan Pol M27
  • 20x / 8 NA Plan-Apochromat M27
  • 40x / 95 NA Plan-Apochromat M27
• Slide racks:
  • Standard microscope slides 75 x 25 mm
  • Macro sections slides 75 x 50 mm
• File formats:
  • “.czi” native lossless or lossy file format with JpegXR compression
  • “BigTiff” after post processing
  • “OME.TIFF” after post processing

Whole Genome Sequencing (WGS) is a comprehensive and high-throughput technique that enables the complete DNA sequence of an organism’s entire genome. WGS is a powerful tool with applications in various fields, including genomics research, personalized medicine, and clinical diagnostics. It provides a comprehensive view of an organism’s genetic makeup, enabling a deeper understanding of genetic variations, evolution, and the genetic basis of diseases.

Bioinformatic analysis of WGS data can be provided as a combined service by the National Facility for Data Handling and Analysis. Please select: SID: NF62.02.01

Whole Exome Sequencing (WES) is a targeted sequencing approach that focuses on sequencing the protein-coding regions of the genome, known as the exome. The exome comprises the exons, which are the coding regions of genes, and represents only a small fraction (about 1-2%) of the entire genome. Despite this, the exome contains most known disease-causing mutations, making WES a cost-effective alternative to Whole Genome Sequencing (WGS) for many applications. WES is widely used in both clinical and research settings. In clinical genetics, it is employed for diagnosing genetic disorders, identifying causative mutations, and understanding the genetic basis of rare diseases. In research, WES is valuable for studying the genetics of complex traits and diseases.

DNA sequencing is critical for genetic research, evolutionary studies, and personalized medicine, where it helps to uncover the genetic basis of diseases, track hereditary conditions, and guide targeted therapies. It provides a detailed understanding of an organism’s complete genetic makeup, offering insights into complex biological processes and evolutionary relationships.

Whole-Genome Sequencing (WGS) involves sequencing the entire genome, including both coding and non-coding regions. WGS provides the most comprehensive view of an organism’s genetic information, as the focus is not only on identifying genetic variants (e.g., single nucleotide variants, insertions, deletions, copy-number variation), but also on identifying rare variants, structural variations, and novel mutations in both coding and non-coding regions. Different algorithms will be applied for germline or somatic samples – the former algorithms are designed to identify inherited variants present in all cells, whereas the latter algorithms focus on detecting mutations acquired in specific tissues which are present only in a subset of cells thus requiring specialized methods to account for tissue purity and heterogeneity.

Exome sequencing is an application of DNA sequencing (see NF62.02.01) that focuses on preferentially sequencing the exons, or protein-coding regions, which make up about 1-2% of the genome and are more likely to harbor disease-causing mutations. It is used to study genetic variations that affect protein function, thus particularly in disease research. The focus is on identifying genetic variants (e.g., single nucleotide variants, insertions, deletions, copy-number variation). Different algorithms will be applied for germline or somatic samples – the former algorithms are designed to identify inherited variants present in all cells, whereas the latter algorithms require specialized methods to account for tissue purity and heterogeneity to detect mutations acquired in specific tissues which are present only in a subset of cells.

This service allows for the creation of web applications and web services that are of interest to the scientific community. Web based applications often represent better solutions compared to desktop applications, the latter entailing manual software installation, software copyright and licensing, software updates, operating systems compatibility, and finally dealing with system requirements. Many of these issues are solved by the adoption of client-server architectures where users can access services hosted on a remote machine. This includes both full-fledged web applications, available through a web browser interface, as well as lower-level services such as APIs.

The Volumetric EM service provides segmentation and analysis of structures in serial EM section data. This service is restricted to segmentation and analysis of a finite number of structures per image (ie, mitochondria, organelles, vesicles, other similar objects of study). The proposal must include specific examples of the structure(s) of interest in order to judge feasibility.  This service includes, but is not necessarily limited to, the following use-cases:

• Data pre-processing: de-streaking, alignment, contrast adjustment.
• Data annotation: generating dense labels covering structures of interest for the purpose of training AI algorithms.
• Model Development: Training and deployment of AI segmentation algorithms specific to the research question.
• Downstream Analysis: Analysis of segmented structures, morphology, number, distribution.

While these are examples of the services we can provide, we anticipate that most projects will require some combination of tools and services and so we will work with successful Applicants to craft pipelines that fulfil their analysis needs, as well as provide training and support in their future use. Our ethos is to work openly and transparently with our Users in the spirit of scientific collaboration. During the application phase, it will only be necessary to describe the data and the desired form of the analysis result; the precise details of the analysis will be discussed with the Applicants upon selection of the project.

This service is designed to cover imaging on large volumes and includes sample preparation by either high-pressure freezing or chemical fixation, freeze substitution, resin embedding, section preparation by ultramicrotomy and S/TEM imaging at 300kV. The User can provide specimen at any stage, so being already fixed, stained, embedded as well as already sectioned and made ready for EM acquisition. A maximum of 2 specimens can be processed per unit of service, and 4 samples maximum (i.e., replicates) can be prepared per specimen. High-pressure freezing will be performed on a Leica EM ICE. Freeze substitution will be performed on a Leica AFS2, and a User might specify a substitution protocol of preference, which will be utilized if compatible with NF practices. Similarly, for chemical fixation, the User is allowed to specify their protocol of preference, which will be adopted if compatible with NF practices. For resin-embedding, the User could provide a resin of preference, or alternatively, the currently available resin be used. Sections of resin-embedded sample will be prepared on Leica UC7. A maximum of 1 resin-embedded sample can be processed for unit of service, to be sectioned in ribbons, and to be applied to a maximum of 4 TEM support grids. User might specify thickness of final sections, and these recommendations will be followed if compatible with the NF practices. User might provide TEM grid support of preference, otherwise the currently available grid support will be used. Imaging will be performed an a Thermo Scientific Spectra 300 kV either in TEM, equipped with CETA 2 camera, or STEM mode in BF/DF/HAADF. Imaging conditions, if requested by the Usermust be compatible with NF practices. Microscope Access is granted for a maximum of 5 days per unit of service, including all steps from sample loading to alignments and collection, and according to NF staff availability. All steps of this service will only be performed by NF staff. To maximise efficient use and Access to high-end TEM instruments, priority will be given to applicants specifically requesting parts of this service (e.g. only sectioning, or only S/TEM imaging of already prepared sections).

Visium HD Spatial Gene Expression (10X Genomics) and the Visium Spatial Gene Expression for Fresh Frozen (Direct placement) allow the spatial profiling of gene expression within intact tissue sections. The protocols allow for the analysis of gene expression while preserving the spatial context of cells within a tissue sample.

Total RNA sequencing is a powerful and widely used molecular biology technique that aims to analyse and quantify the entire transcriptome of a biological sample. The transcriptome represents the complete set of RNA molecules, including messenger RNA (mRNA), and non-coding RNAs, present in a cell or tissue. Total RNA sequencing provides a comprehensive view of the transcriptome, allowing researchers to gain insights into gene expression patterns, identify novel transcripts, and understand the regulatory mechanisms underlying various biological processes.

Our service focuses on introducing INDELs (frame-shift mutations) to generate gene knock-out lines (KO), each targeting a single gene in either hPSCs or immortalized/cancer cell lines.

Characterization of each engineered cell line includes:

• Cell identity confirmation using STR analysis
• Confirmation of desired editing via Sanger sequencing.

• Karyotyping (Q-banding) and identification of Copy Number Variations (CNVs) at high resolution
• Master bank post-thaw viability and mycoplasma testing.
• Optional evaluation of undifferentiated stem cell markers and pluripotency markers upon 3-germ layer differentiation assay.

We accept cell lines in a cryopreserved state, with a minimum of 1 x 10^6 cells per cryovial.

We generate up to 3 homozygous knockout (KO) clones. Heterozygous KO clones can be provided upon agreement with the applicant. If technically feasible, clones will be analyzed for the occurrence of allelic dropout.

The service typically requires 2 to 3 months for completion and includes the delivery of 1-3 clones, each provided in 10 cryopreserved vials.

The Visium Spatial Gene Expression solution from 10X Genomics enables spatial profiling of gene expression within intact tissue sections. This protocol allows for the analysis of gene expression while preserving the spatial context of cells within a tissue sample. It provides valuable insights into spatially distinct gene expression patterns and cell-type localization, facilitating a deeper understanding of tissue organization, disease progression, and cellular microenvironments.

Small RNA sequencing is a specialized technique designed to analyse and profile small RNA molecules present in a biological sample. Small RNAs are short RNA molecules, typically ranging from 18 to 30 nucleotides in length, and they play essential roles in various cellular processes, including gene regulation, RNA silencing, and post-transcriptional control. Small RNA sequencing is widely used to study the expression profiles of miRNAs and other small RNAs, providing valuable insights into their roles in gene regulation, development, and disease.

Single-cell multiome ATAC + Gene Expression is a cutting-edge technology that enables the simultaneous profiling of chromatin accessibility and gene expression at the single-cell level, providing a comprehensive view of the molecular landscape within individual cells. This technology is widely used in various biological research areas, including understanding cellular diversity in tissues, identifying cell types and states, deciphering regulatory networks, and gaining insights into how chromatin accessibility relates to gene expression at the single-cell level.

Single-cell multiome sequencing (scRNA-seq + scATAC-seq) is molecular biology technique used to analyze both gene expression and chromatin accessibility of individual cells/nuclei at a high resolution, allowing the resolution of cellular heterogeneity within a biological sample.

Unlike bulk RNA and ATAC sequencing, which averages gene expression and chromatin accessibility across many cells, multiome sequencing enables the identification of distinct cell types, states, and subpopulations both in terms of transcriptional and epigenetic profiles. This technology is particularly useful in studying various processes and biological mechanisms including developmental processes, tumorigenesis, and immunological memory establishment. It is widely applied in biomedical research to advance personalized medicine, immunology, cancer research, and tissue regeneration studies.

Multiome datasets include two different assays: gene expression (GEX) and chromatin accessibility (ATAC).

Single-cell immune profiling with 10x Genomics technology is a powerful method for 5’ RNA sequencing at the single-cell level and to profile at the same time the T-cells and/or B-cells receptors at single cell level by sequencing the V(D)J regions.

Single-cell immune profiling-V(D)J is a powerful molecular biology technique used to profile both 5’ gene expression and T-cell and/or B-cell receptors of individual cells at a high resolution allowing the characterization of cellular heterogeneity and clonal expansion within a biological sample.

Unlike bulk RNA and T/B-cell receptor (TCR/BCR) sequencing, which allow to study gene expression and TCR/BCR repertoires across many cells, single-cell immune profiling-V(D)J enables the identification of distinct cell types, states, and subpopulations both in terms of transcriptional profile (GEX data) and TCR/BCR repertoires (V(D)J data). This approach is crucial for understanding complex tissues, developmental progression, tumorigenesis, and tracking clonal expansion and immune responses. It is widely applied in biomedical research to advance personalized medicine, immunology, cancer immunotherapy, autoimmune disease and infection disease.

The Single-cell Immune profiling-V(D)J datasets include two modalities: gene expression (GEX) and TCR/BCR (V(D)J).

Single-cell ATAC sequencing with the 10X Genomics platform involves profiling the chromatin accessibility of individual cells/nuclei at a high resolution. The assay, commonly known as scATAC-seq (Single-cell Assay for Transposase-Accessible Chromatin sequencing), utilizes the 10X Genomics Chromium system to barcode and index individual nuclei, it provides valuable insights into the epigenomic landscape of individual cells, allowing researchers to study cellular heterogeneity and regulatory processes at high resolution.

Single-cell 3′ RNA sequencing with 10x Genomics technology is a powerful method for studying gene expression at the single-cell level. It enables the profiling of thousands to tens of thousands of single cells in parallel. It captures heterogeneity within cell populations, allowing the identification of rare cell types and subpopulations. It reveals differences in gene expression between individual cells, providing a more nuanced understanding of cellular diversity.

The Fixed RNA Profiling assay, also called the Single Cell Gene Expression Flex assay by 10X Genomics, is a way to prepare single cell RNA-seq libraries from formaldehyde-fixed cells and tissues as well as FFPE tissue blocks (please consider that from FFPE tissues the isolated population primarily consists of nuclei suspension, thus the RNA-seq analysis is to be intended at nuclear level). This process allows researchers to lock in the biological state of their samples at the time of fixation and store the fixed cell suspension for at least 6 months at -80°C.

The National Facility for Genomics will provide sequencing of pools of libraries prepared by the users with the NovaSeq6000 sequencing platform (Illumina).

The NovaSeq 6000 is a high-throughput sequencing platform developed by Illumina. It’s designed to handle a wide range of applications and large-scale genomic projects.

NovaSeq 6000 is commonly used for: Whole Genome Sequencing (WGS), Exome Sequencing (WES), Transcriptome Analysis (bulk RNA-Seq or single-cell RNAseq), Metagenomics, Epigenomics studies, Population Genomics.

The National Facility for Genomics will provide sequencing of pools of libraries prepared by the users with the NextSeq2000 sequencing platform (Illumina).

The NextSeq 2000 is a next-generation sequencing platform developed by Illumina. It is designed to offer high-throughput sequencing with flexibility for various applications.

The NextSeq 2000 is commonly used for: Whole Genome Sequencing (WGS), RNA Sequencing (RNA-Seq and single-cell RNA-seq), Targeted Sequencing (amplicon sequencing), Exome Sequencing (WES), Metagenomics, ChIP-Seq and Epigenomics.

The National Facility for Genomics will provide sequencing of pools of libraries prepared by the users with the MiSeq sequencing platform (Illumina).

the MiSeq is a benchtop next-generation sequencing platform developed by Illumina. It’s designed for smaller-scale sequencing projects and offers flexibility for various applications.

MiSeq is commonly used for: Targeted Sequencing (amplicon sequencing), Small Genomes Sequencing, Metagenomics, 16S rRNA Sequencing, Small RNA-Seq, Viral genomes Sequencing.

Single-cell RNA sequencing (scRNA-seq) is a technique used to analyse gene expression at the individual cell level, making it possible to resolve cellular heterogeneity within a biological sample. Unlike bulk RNA sequencing, which averages gene expression across many cells, scRNA-seq enables the identification of distinct cell types, states, and subpopulations.

This approach is crucial for understanding complex tissues, developmental processes, and disease progression, as it reveals how gene expression varies from cell to cell, and it is widely applied in biomedical research to advance personalized medicine, immunology, cancer research, and tissue regeneration studies.

Single-cell ATAC sequencing (scATAC-seq) is a powerful molecular biology technique used to profile the chromatin accessibility of individual cells/nuclei at a high resolution. Chromatin accessibility refers to the degree to which DNA within chromatin is accessible by cellular machinery, particularly those parts involved in transcription, such as transcription factors and RNA polymerase.

Unlike bulk ATAC sequencing, which cannot determine the chromatin states of individual subpopulations of cells within a sample, scATAC-seq is widely used to provide valuable insights into chromatin accessibility, transcription factor binding, epigenetic modifications, and gene regulation. This technology is particularly useful in studying various processes and biological mechanisms including developmental processes, tumorigenesis, and immunological memory establishment.

Many workflows available in other Infrastructural Units of the National Facility for Structural Biology may require a rapid sample quality control verification (if not performed by the users) or a step of sample optimisation, for instance via size-exclusion chromatography just prior to experiments. These services, which differ from the full characterisation of the macromolecular sample because of their ancillary role with respect to other IU workflows, may only be requested in combination with such workflows.

iPSCs are generated from Peripheral Blood Mononuclear Cells (PBMCs) using footprint-free reprogramming Sendai virus technology.

Every iPSC line undergoes thorough testing to ensure quality and consistency:

• Cell identity confirmation using STR analysis
• Sendai virus clearance
• Expression of undifferentiated stem cell markers
• Assessment of pluripotency markers through 3-germ layer differentiation assay
• Karyotyping (Q-banding) and identification of Copy Number Variations (CNVs) at high resolution
• Master bank post-thaw viability and mycoplasma testing.

Both live and frozen purified PBMCs qualify for the service.

The service typically requires 3 to 4 months for completion and includes the delivery of 1-3 clones, each provided in 10 cryopreserved vials.

Fibroblasts are reprogrammed using footprint-free, non-modified RNAs (nm-RNA).

Every iPSC line undergoes thorough testing to ensure quality and consistency:

• Cell identity confirmation using STR analysis
• Sendai virus clearance
• Expression of undifferentiated stem cell markers
• Assessment of pluripotency markers upon 3-germ layer differentiation assay
• Karyotyping (Q-banding) and identification of Copy Number Variations (CNVs) at high resolution
• Master bank post-thaw viability and mycoplasma testing.

Both live and frozen fibroblasts at a low passage number (maximum passages 5) qualify for the service.

The service typically requires 3 to 4 months for completion and includes the delivery of 1-3 clones, each provided in 10 cryopreserved vials.

This service comprises 8 hours of equipment time on the Orbitrap Astral LC-MS. The configuration of the system is: Vanquish NEO HPLC in trap and elute configuration. Mobile phase A is 0.1% formic acid in water, mobile phase B is 80% acetonitrile in water with 0.1% formic acid. The MS is set up with an EasySpray source. The MS is operated for high- throughput, with EasySpray pepMap columns (15cm, 150µm diameter, 10nm pore size). It is possible to use the FAIMS filter to improve sensitivity or fractionate samples online. Overhead on measurements due to sample loading and column washing is about 2 minutes per injection. Typical gradient length 15 minutes (no gains are typically observed beyond this gradient length due to the speed of the Astral analyser and the operation in high flow). This configuration allows for up to 100 samples per day. Both DIA and DDA proteomics workflows are available.  The service does not comprise sample preparation from proteins to peptides.

This service comprises 8 hours of equipment time on the Orbitrap Astral LC-MS. The configuration of the system is: Vanquish NEO HPLC in trap and elute configuration. Mobile phase A is 0.1% formic acid in water, mobile phase B is 80% acetonitrile in water with 0.1% formic acid. The MS is set up with an EasySpray source. The MS is operated for high- load, using EasySpray pepMap NEO columns (50cm, 75µm diameter, 10nm pore size). It is possible to use the FAIMS filter to improve sensitivity or fractionate samples online. Overhead on measurements due to sample loading and column washing is about 10 minutes per injection. Typical gradient length 30 minutes (no gains are typically observed beyond this gradient length due to the speed of the Astral analyser). Both DIA and DDA proteomics workflows are available. Each proposal can request up to 9 sessions for a total of 3 days of MS time. The service does not comprise sample preparation from proteins to peptides.

We offer access to expert support in isolating and purifying proteins from a variety of biological sources. We offer customisable workflows, including cell lysis via sonication or homogenization, depending on any given sample’s specific requirements. The service can accommodate both purification from standard small and medium-volume lysates, or isolation from large-volume sample preparations, ensuring flexibility for different experimental needs. Our purification techniques include affinity, ion exchange, and size-exclusion chromatography, delivering (when possible) high-quality proteins suitable for downstream applications such as structural analysis or functional assays.

The service offers dedicated pipelines for small- and large-scale cell cultures and expressions:

• Small scale: 15 litres fermenter useful for initial trial test before moving to larger scale fermenter or small expression using both bacteria and yeast cells. Typical culturing strategies are available such as batch and fed batch (up to 2 feeds). Max working volume of 10 litres.
• Large scale: 150 litres fermenter dedicated to large cultivation and protein production. Typical cultivation strategies are available, including batch and fed batch (up to 2 feeds). Max working volume of 100 litres.

In both cases, cells will be collected and separated from the supernatant using dedicated superspeed centrifuges or a High-Speed Tubular Centrifuge. Upon request, the cells can be lysed using a continuous flow cell disruptor or by a high-capacity cryogenic grinder.

The service offers dedicated pipelines for small- and large-scale cell cultivation and protein expressions:

• Small scale: utilizing commercial Expi293F cells, transient transfection is conducted with a plasmid containing the gene/s of interest (potentially with a fluorescent marker), provided by the applicant. Cell cultivation and transfection take place in Erlenmeyer flasks within dedicated shaker incubators; typically, 500 ml of cell culture per 2-liter flask is utilized up to a total cultivation volume of approximatively 10 litres. Transfection of an Expi293F cell culture is performed at approximately 2 to 3 x10^6 cells/ml, following a modified protocol with polyethylenimine (PEI). While no expression test will be performed, the efficiency of transfection can be readily assessed if the plasmid includes a fluorescent marker. Transfected cells are subsequently harvested by centrifugation, and either the frozen pellet or the refrigerated supernatant will be provided.
• Large scale: bioreactor cultivation and protein expression are performed either with commercial cell lines (e.g., Expi293F cells) transiently transfected with applicant-provided plasmids, or with any mammalian cell line adapted to suspension cultivation supplied by the applicant. Preliminary tests are normally performed using a 0.5 litres scale bioreactor to define and optimize bioreactor cultivation conditions. Subsequently, a 10 litres scale bioreactor enables large-scale cell cultivation and protein expression, employing both transient transfection protocols and stable cell line expressions. Moreover, the implementation of perfusion, utilizing an acoustic retention device, may be explored to increase the biomass production and, consequently, protein expression. Similar to small-scale operations, no expression test will be performed. Instead, monitoring of transfection efficiency for transient expression can be undertaken if the plasmid carries a fluorescent marker or fluorescently tagged protein of interest.

The service aims to cover the entire process of Baculovirus protein expression, starting from a Bacmid. Initially, commercial Sf9 cells are transfected with the Bacmid, supplied by the applicant, which contains the gene of interest and a fluorescence marker. The initial stage results in a low-titre Baculovirus suspension (P1 generation), followed by a second amplification (P2 generation) to produce a high-titre suspension. The P2 Baculovirus suspension could be directly used for protein expression or another round of Baculovirus amplification is performed before infecting commercial High Five insect cells (at 0.5×10^6 cells/ml). The virus amount used depends on the construct and might need optimization in initial small-scale infections. Protein expression is monitored indirectly by checking for the fluorescence marker (carried by the Bacmid) and cell viability. Finally, cells are harvested via centrifugation, and the pellet or supernatant is made available.

This service provides support for the code maintenance lifecycle for scientific software which has already reached a sufficiently mature development stage. The general aim is to provide best tools and practices to achieve better quality of code, software reproducibility and efficiency, making it a high value product and ensuring its long-term survival.

The service focuses on two main areas:

1. Containerization, standardization and improvement of bioinformatics pipelines;
2. Maintenance of general software for the life sciences.

The first area involves procedures to make existing bioinformatics pipelines more efficient and reproducible. Additional features such as data reporting can also be developed. The target output is a Nextflow pipeline composed of containerized modules, with a focus on code execution efficiency and reproducibility.

The second area encompasses the broader category of software that needs to be improved and/or updated to avoid obsolescence.

Nikon Ti2 inverted microscope with CrestOptics X-light V3 spinning disk scan head.

• Available lasers sources: 405 nm, 446 nm, 477 nm, 520 nm, 547 nm, 638 nm, 749 nm
• Available laser sources (TIRF and FRAP): 405 nm, 488 nm, 561 nm, 640 nm
• Four high-speed, back sided illuminated sCMOS cameras (Prime 95B, 25 mm FOV)
• X-light V3 spinning disk scan head (50 µm pinhole disk)
• Incubator (CO₂, temperature)
• TIRF condenser
• Photomanipulation module
• NIS elements software
• Available objectives:
• 4x/0.20 Plan Apo
• 10x/0.45 lD CFI Plan Apochromat
• 20x/0.8 lD CFI Plan Apochromat
• 20x/0.45 S Plan Fluor ELWD
• 40x/1.25 Sil lD CFI Plan Apochromat
• 100x/1.35 Sil lS CFI SR HP Plan Apochromat
• 100x/1.49 Oil CFI Apo TIRF

We offer sample preparation by negative stain EM followed by TEM imaging at 120 kV. A maximum of 4 service units (i.e. 4 days of screening and sample preparation on 1 specimen or 4 different ones) can be requested. For each session a maximum of 8 grids can be prepared and processed. Imaging will be provided for a maximum of 8 continuous hours per unit of service. An optional “polishing” size-exclusion chromatography (SEC) step can be performed on thawed material by the Biophysics Unit, as and if considered necessary by NF Staff. 400 mesh copper grids with amorphous carbon film layer will be used as support. Glow discharging will be performed by a Pelco EasyGlow device. Staining will be performed with a 2% (w/v) uranyl acetate aqueous solution. Imaging in TEM mode at 120 kV will be performed on a Thermo Scientific Talos L120C equipped with CETA 16M camera. Imaging conditions requested by the User, if provided, must be compatible with NF practices. This service will only be performed by NF staff.

Nanopore sequencing of small gDNAs will be performed with the native barcoding kit that refers to a set of reagents and protocols designed to enable the simultaneous sequencing of multiple samples by adding unique barcodes to each sample before sequencing. This approach is particularly useful for studying small bacteria genomes (gDNA) as it allows for high-throughput sequencing and analysis of multiple samples using a single flowcell.

This approach provides a cost-effective and efficient way to sequence multiple small bacteria genomes simultaneously, making it a valuable tool in microbiome studies, environmental monitoring, and other applications in microbial genomics.

Nanopore sequencing is a next-generation sequencing technology that uses nanopores to directly sequence DNA molecules and to analyse DNA bases modifications. This method is known for producing long reads, and in some cases, ultra-long reads, making it valuable for various genomic applications.

Direct RNA Sequencing with Nanopore technology is a cutting-edge method for sequencing RNA molecules without the need for conversion to complementary DNA (cDNA) as required in traditional RNA sequencing methods. The protocol involves the direct sequencing of RNA strands through nanopores, allowing for the real-time detection of RNA sequences and RNA bases modifications. Direct RNA Sequencing with Nanopore technology offers the advantage of studying RNA molecules in their native state, providing valuable insights into RNA processing, alternative splicing, and modifications without the need for reverse transcription. It is particularly valuable for capturing the full complexity of the transcriptome.

Cell-free DNA (cfDNA) sequencing with Nanopore technology represents a revolutionary approach to interrogate the genetic information present in circulating DNA, often extracted from blood plasma. Unlike traditional sequencing methods, this protocol allows for the direct sequencing of cfDNA without the need for intermediate steps such as PCR amplification or conversion and allows the analysis in real time of the methylation status of circulating cfDNA. Its applications extend to clinical diagnostics, providing valuable information for personalized medicine and disease monitoring such as liquid biopsy for cancer detection, monitoring treatment response, and identifying minimal residual disease.

Nanopore cDNA sequencing is a powerful technique that combines the benefits of nanopore sequencing technology with the study of complementary DNA (cDNA), which represents the transcribed RNA in a biological sample. This method allows researchers to investigate gene expression, alternative splicing, and other aspects of RNA biology with long-read sequencing capabilities. Nanopore cDNA sequencing offers several advantages, including the ability to generate long reads that span entire transcripts. This makes it particularly valuable for studying complex transcriptomes, characterizing novel isoforms, and exploring the dynamics of gene expression in various biological contexts.

Small RNA sequencing is a specialized technique designed to analyze and profile small RNA molecules present in a biological sample. It is widely used to study the expression profiles of miRNAs and other small RNAs, providing valuable insights into their roles in gene regulation, development, and disease. Small RNAs are polymeric ribonucleic acid molecules with a length lower than 200 nucleotides, comprising microRNA (miRNA), PIWI-interacting RNA (piRNA), small interfering RNA (siRNA), and tRNA-derived small RNA (tsRNA).

miRNAs are the most studied type of small RNAs, constituted by 20 to 25 nucleotides. They participate in several processes and can regulate gene expression at a posttranscriptional level. miRNAs can also act as transcription factors by binding the seed sequence within 3’UTR of target genes, leading to a variety of cell activities at different levels.

High resolution, functional imaging based on high density MEA electronic interfaces measuring extracellular voltage potentials. The system available is the multichannel amplifier BioCAM DupleX and planar MEAs electronic chips equipped with 4,096 electrodes (3BRAIN AG, Pfäffikon SZ, Switzerland). The MEAs chip is the CorePlate^TM 1W38/60 (NF55.004_MEA chip testing and provision) with a recording area of 3.8 mm x 3.8 mm, each electrode is 21 mm x 21 mm with a 60 mm pitch. The assay is suitable for the characterization of functional phenotypes in cell networks, and for compound screening. The high density of electrodes allows to collect a large amount of data within a single recording, and it is particularly suitable for accessing the electrical phenotype of brain and cardiac organoids.

Microbiome analysis using 16S and ITS amplicon sequencing is a widely used technique to study the composition and diversity of microbial communities, particularly bacteria and fungi. The 16S ribosomal RNA (rRNA) gene is a molecular marker found in the genomes of bacteria and archaea, and its variable regions are commonly used for taxonomic classification, while ITS is used to profile fungal communities.

Microbiome analysis using 16S and ITS amplicon sequencing is valuable in a range of fields, including environmental science, human health, and agriculture. It provides a cost-effective way to characterize microbial communities and understand their roles in various ecosystems or host-associated environments.

Methylation sequencing (Methyl-seq) is a technique used to study DNA methylation, a key epigenetic modification where methyl groups are added to DNA molecules, typically at cytosine bases in CpG sites. This method allows researchers to identify and quantify methylation patterns across the genome, providing insights into gene regulation, cellular differentiation, development, and disease processes such as cancer, where abnormal methylation often occurs.

The Biophysics Unit is equipped with instruments designed to determine the strength and biophysical properties of macromolecular or protein-ligand interactions, including association and dissociation constants, as well as stoichiometry of binding. Available techniques include isothermal titration calorimetry, microscale thermophoresis, and bio-layer interferometry (which can also be utilized for quantifying known components in complex mixtures).

Light microscopy analysis encompasses the analysis of data generated by any light microscopy modality (ie, brightfield, phase contrast, widefield epi-fluorescence, confocal, lightsheet, etc) and across any sample type.

The services we provide include, but are not necessarily limited to, the following use-cases:

• Image restoration and denoising: Removal of pixel-independent noise from images to increase signal-to-noise ratio (SNR).
• Semantic and Instance segmentation: Identification and segmentation of objects in an image, generation of image masks.
• Quantitative Image Analysis: Quantification of intensity levels in images or segmented objects.
• Morphometric Analysis: Analysis of shape and morphology of segmented objects.
• Custom pipeline development: Construction of an analysis pipeline combining two or more individual steps.

While these are examples of the services we can provide, we anticipate that most projects will require some combination of tools and services and so we will work with users to craft pipelines that fulfil their analysis needs, as well as provide training and support in their future use. Our ethos is to work openly and transparently with our users in the spirit of scientific collaboration. During the application phase, it will only be necessary to describe the data and the desired form of the analysis result; the precise details of the analysis will be discussed with the users upon selection of the project.

Leica Thunder motorized epifluorescence microscope with integrated digital clearing.

• Available LEDs: 395 nm, 438 nm, 475 nm, 551 nm, 555 nm, 575 nm, 635 nm and 730 nm
• Incubator (CO₂, temperature)
• Leica K8 sCMOS camera
• Leica LAX software
• Leica Navigator
• Computational Clearing module
• Available objectives
• 4x/0.10 HI PLAN
• 10x/0.32 HC PL FLUOTAR PH1
• 20x/0.40 Korr HC PL FLUOTAR L PH1
• 40x/0.60 Korr HC PL FLUOTAR PH2
• 63x/1.40-0.60 HC PL APO Oil

Leica Stellaris 8 confocal microscope with tandem scanners (resonant and conventional galvanometer scanner), STED and FALCON modules.

• Available lasers sources: 405 nm laser and tunable white light laser with up to 8 simultaneous lines between 440 nm and 790 nm
• 4 hybrid detectors
• STED module with 775 nm depletion laser
• FALCON module for fluorescence lifetime imaging
• Incubator (CO₂, temperature)
• Leica LAX software
• Leica Navigator
• Available objectives:
• 10x/0.40 HC PL APO CS2
• 20x/0.75 HC PL APO CS2
• 40x/1.30 HC PL APO Oil CS2
• 63x/1.40 HC PL APO Oil CS2
• 100x/1.40 HC PL APO Oil STED

Leica Stellaris 8 confocal microscope with tandem scanners (resonant and conventional galvanometer scanner), incubator and FALCON module.

• Available lasers sources: 405 nm laser and tunable white laser with up to 8 simultaneous lines between 440 nm and 790 nm
• 4 hybrid detectors
• Incubator (CO₂, temperature)
• FALCON module for fluorescence lifetime imaging
• Leica LAX software
• Leica Navigator
• Available objectives:
• 10x/0.40 HC PL APO CS2
• 20x/0.75 HC PL APO CS2
• 40x/1.30 HC PL APO Oil CS2
• 63x/1.40 HC PL APO Oil CS2

If the cell is supplied with a donor DNA template, Cas9-induced DSBs can be repaired by integrating DNA sequences of various lengths using homology directed repair (HDR) mechanism. We provide Knock-In (KI) services to obtain reporter cell lines (including safe harbor integrations), edited cells harboring specific single nucleotide corrections as well as labelled proteins (e.g., with fluorescence or a protein tag).

Characterization of each engineered cell line includes:

• Cell identity confirmation using STR analysis
• Confirmation of desired editing via Sanger sequencing.

• Karyotyping (Q-banding) and identification of Copy Number Variations (CNVs) at high resolution
• Master bank post-thaw viability and mycoplasma testing.
• Optional evaluation of undifferentiated stem cell markers and pluripotency markers upon 3-germ layer differentiation assay.

We accept cell lines in a cryopreserved state, with a minimum of 1 x 10^6 cells per cryovial.

The service typically requires 2 to 4 months for completion and includes the delivery of 1-3 clones, each provided in 10 cryopreserved vials.

Fluorescence-based time-lapse recordings of intracellular ion oscillations, based on confocal or epifluorescence microscopy.

• Imaging on Nikon Ti2 spinning disk with four cameras, TIRF and FRAP module (SID: 007 or equivalent system suggested by the NF staff).

The Structural Proteomics unit will perform integrative structural modelling combining medium/ low resolution (from 7 to 30 Å) cryo-EM densities and crosslinking MS data acquired at our NF. In case of very large systems, negative stain data may also be used. This does not refer to model building in cryo-EM densities, but to calculations of localization of subunits or of areas not observed in EM maps. This task can be performed on data from services SB-IU4-A or SB-IU4-B only. Example software: DisVis, integrative modelling platform (IMP), AlphaLink/ AlphaLink2. This service will be performed by NF staff, but training will be available if requested, provided basic bioinformatics skills of the User (bash terminal, python).

This service provides high-resolution cryo-TEM data collection of vitrified specimens. To ensure efficient usage of high-end microscope time this service is exclusively dedicated to EM grid ready for data collection at the time of application (i.e. user cannot request this service together with service SB-IU1-B – Cryo-EM Screening). A maximum of 2 samples for data collection can be requested per application. For each sample a maximum of 48hr of microscope time can be requested per application. According to instrument availability and experimental needs, data collection will be carried out either on 200kV or 300kV microscope systems. The User can provide cryo-TEM grids either unmounted or mounted on a Thermo Scientific cartridge. In case of an unmounted grid, clipping of the specimen in Thermo Scientific cartridges will be performed by NF staff. In case of User-provided already-clipped grids, these will be inspected by NF staff prior to acceptance. Imaging at 200 kV will be performed on a Thermo Scientific Glacios while imaging at 300 kV will be performed on a Thermo Scientific Titan Krios G4, both equipped with a Falcon 4i direct electron detector and a Selectris X energy filter. Imaging conditions (i.e., dose, pixel size, magnification, etc.), if requested by the User, must be compatible with the NF best practices. Microscope time includes all steps from clipping to loading and TEM alignments and according to NF staff availability. For single-particle acquisition, User might opt for beam-image shift assisted data collection (~ 450 – 600 movies/hour) or stage movement for each hole (~ 100 – 250 movies/hour). This service will only be performed by NF staff.

The GeoMx Digital Spatial Profiling service provides cutting-edge technology for spatially resolved gene expression and protein analysis. This service supports fresh-frozen (FF), fixed-frozen and formalin-fixed, paraffin-embedded (FFPE) tissue samples, enabling researchers to explore spatial biology in diverse sample types.

User consultation is an essential first step in assuring high-quality flow cytometric data. Our staff are available to guide new and current Users to meet each projects’ needs, addressing all critical steps between the conception of the flow cytometry experiment, the appropriate steps for sample preparation including the required experimental controls, the acquisition of samples and the analysis of the data generated.

Full-service sorting of rare populations from heterogeneous samples, cell cloning (single cell deposition into multi-well plates), particle enrichment, and high purity bulk sorts.

High-recovery and indexed single-cell sorting for sequencing. Cell sorting of many cell types including:

• immune cell and hematopoietic stem cell subsets
• mesenchymal stem cells
• viable cytokine producing cells
• general cell sorting approaches for cell lines
• transfected cells, including iPSCs Sorting Technical Details

Capable of standard and high-speed sorting of up to 6 populations simultaneously. Sorted cells may be recovered in numerous devices including:

• 6, 12, 24, 28, 96, 384, 1536 well plates & 96 deep well plates
• 2, 0.5, 1.2, 1.5, 2, 5 mL tubes for 6-way sorting
• 15, 50 mL tubes for 2- or 3-way sorting
• Slides or Ibidi vessels
• Custom vessels may also be

The sorter is equipped with 6 lasers having the following emissions: 355 nm 405 nm, 488 nm, 560 nm, 592 nm, 645 nm.

Proper controls for each session must be included with the analysis samples. For example:

• Unstained, unlabeled or other cellular controls
• Spectral overlap compensation controls
• FMO/FMX controls when appropriate

Autonomous Flow Cytometry Analysis (after completion of a training session) or Operator Assisted Flow Cytometry Analysis.

The following are examples of analyses we offer:

• Prepared immune cell and hematopoietic stem cell
• Prepared and labelled samples of mesenchymal stem
• Analysis of cytokine producing
• Multi-color extracellular and intracellular stained
• DNA content analysis (single color or multi-color).
• Cell proliferation
• Apoptosis
• Functional and metabolic assays (mitochondrial function, ROS production, lipid metabolism).
• Analysis of bead assays (e.g., Cytokine/chemokine bead assays). Analyzer Technical Details:
• Integrated absolute count
• Plate loader supporting 96 wells plate and 96 deep wells U, V or Flat bottom
• Different sample injection modes, manual or automatic
• Wide sample flow rate and minimal dead sample volume (about 20 µl)

Analyzers are equipped with 5 or 6 lasers having the following emissions: 355 nm, 405 nm, 488 nm, 561 nm, 640 nm and 808 nm. SSC parameter is available for the 488 and the 405 nm lasers lines.

• Capable of complex polychromatic
• Advice in experimental design and dedicated training sessions are also available. Proper controls for each session must be included with the analysis For example:
• Unstained, unlabeled or other cellular
• Spectral overlap compensation
• FMO/FMX controls when

This service is dedicated to projects where specific single molecule assays have been already established or, alternatively, should be combined with ‘assay development’ service as a simple ‘package’. Examples of experiments currently performed routinely by the unit: (i) detecting changes to DNA structure upon protein binding e.g., melting, looping, compaction; (ii) characterizing protein behaviour upon binding to DNA (diffusion, direct motion, interactions with other proteins). This service can be performed by the NF staff or by a trained User. The training can be provided by the unit as part of this service. Full training typically requires 3-5 days. The User can spend up to 10 days at the NF. The work can be split into two visits. Original data will be handed to the User as .h5 files. The unit will assist the User in Accessing the files using Pylake. In case of kymographs, NF staff can provide a short training on existing tools such as Lakeview.

Development and execution of custom gene editing projects.

This service is designed to offer expertise and technical support for the implementation of genome engineering projects that demand customized approaches not presently covered by off-the-shelf knock-out and knock-in editing services.

The proposal should provide a thorough explanation of the rational, goals, and expected outcomes to facilitate a comprehensive evaluation.

Tailored gene editing projects involve completing sequential checkpoints before moving on to the implementation of the proposed experiment (Phase 3). Throughout these phases, significant interaction with the applicant is expected.

• Phase 1 (Strategy design): A tailored strategy will be designed following preliminary meetings and discussed with the applicant before proceeding to the next phase.
• Phase 2 (Strategy validation): The designed approach will undergo testing in the relevant cell line to assess its effectiveness (efficiency, locus accessibility, …).
• Phase 3 (Target editing generation): Upon successful completion of Phase 2, the validated approach will be applied to generate the desired cell line.

This service provides cryo-Fluorescence Microscopy imaging including sample preparation by plunge-freezing and grids clipping. A maximum of 2 specimens can be processed per unit of service, maximum 4 grids (i.e. replicates) can be prepared per specimen. Glow discharging will be performed by either a Pelco EasyGlow, a Quorum GloQube, or Solarus II plasma cleaner device. Plunge-freezing will be performed on a Thermo Scientific Vitrobot Mk IV or a Leica EM GP2. Applicants may provide TEM grid supports of preference, otherwise the NF staff will use the available TEM grid supports. Widefield imaging will be performed on a Leica Thunder cryo-CLEM system. Confocal imaging will be performed on a Leica Stellaris 5 cryo-CLEM system equipped with white light laser. Both microscopes are equipped with a 50x / 0.9 NA lens. This service is provided for a maximum of 8 continuous hours per unit of service, including plunging, clipping, and imaging. In case of User-provided already-clipped grids (in Thermo Scientific cartridges), these will be inspected by NF staff prior to acceptance. This service will only be performed by NF staff.

This service provides sample preparation by plunge-freezing, grid clipping for autoloader system and cryo-TEM imaging at 200 kV. User can either provide the purified sample in solution, the sample already vitrified on EM grid, or the vitrified sample on an already clipped (i.e. Thermo Scientific autoloader compatible) EM grid. User might request cryo-EM screening on a maximum of 4 different samples per application. For each sample, a maximum of 3 Cryo-EM Screening sessions can be allocated. For each Cryo-EM screening session a maximum of 8 grids can be prepared and processed. An optional “polishing” SEC step can be performed on thawed material by the Biophysics Unit, as and if considered necessary by NF Staff. Glow discharging will be performed by either a Pelco EasyGlow, a Quorum GloQube, or Solarus II plasma cleaner device. Plunge-freezing will be performed on a Thermo Scientific Vitrobot Mk IV or a Leica EM GP2. Applicants may provide TEM grid supports of preference, otherwise NF staff will use holey TEM grid supports available. Imaging will be provided for a maximum of 8 continuous hours per unit of service. In case of grids showing optimal particle distribution and ice quality, this service can be extended for overnight data collection, if compatible with NF staff working hours. Cryo-TEM imaging will be performed only in EF-TEM mode at 200 kV on a Thermo Scientific Glacios equipped with a Falcon 4i direct electron detector and Selectris X energy filter. Imaging conditions requested by the User, if provided, must be compatible with NF practices. In case of User-provided already-clipped grids, these will be inspected by NF staff prior to acceptance. This service will only be performed by NF staff. This service does not cover for buffer screening (i.e. detergents, pH, salt, additives screening or else) or any other grid modifications (i.e. graphene oxide, pegylation, affinity grids or else).

Cryo-Electron microscopy analysis encompasses the analysis of cryo-electron microscopy data, both single particle and tomographic reconstruction. This service includes, but is not necessarily limited to, the following use-cases:

• Single-particle analysis (SPA): Development of image processing pipelines for the reconstruction of single particle 3D density maps, starting from cryoEM raw datasets or pre-processed micrographs/particles. Map validation.
• Atomic Model Building: De novo model building from reconstructed 3D density maps, fitting of existing atomic structures and refining of atomic models. Model validation.
• Analysis of Flexibility and Heterogeneity: Development of image processing pipelines for local reconstruction and refinement of flexible regions and evaluation of the conformational heterogeneity landscape of the macromolecules.
• Tomography reconstruction: Development of image processing pipelines for the reconstruction and analysis of tomograms, starting from tilt-series containing fiducial markers or fiducial less. Segmentation of the tomograms and sub-tomogram averaging (STA).
• Custom pipeline development: Construction of a computational pipeline combining two or more individual steps.

While these are examples of the services we can provide, we anticipate that most projects will require some combination of tools and services and so we will work with successful Applicants to craft pipelines that fulfil their analysis needs, as well as provide training and support in their future use. Our ethos is to work openly and transparently with our Users in the spirit of scientific collaboration. During the application phase, it will only be necessary to describe the data and the desired form of the analysis result; the precise details of the analysis will be discussed with the Applicants upon selection of the project.

The Structural Proteomics unit will perform crosslinking MS on a purified protein complex to identify protein-protein interactions and map residue distances. This service does not include preliminary crosslinking reaction optimisation, which should be performed by the User at their own institution (in consultation with NF staff). This service will only be performed by NF staff.

The Structural Proteomics unit will perform crosslinking MS on a purified protein complex to identify protein-protein interactions and residue distances. This service includes preliminary crosslinking reaction optimisation and proteomics acquisitions. These can be performed by our NF staff provided the protein sample, or by a visiting User with our assistance.

The Structural Proteomics unit will perform crosslinking MS to characterize the interactome of a particular target protein in situ. Crosslinking reactions can be performed on cells, in lysate or after competitive or otherwise non-denaturing (native) elution. As this experiment requires a careful and at times tricky design, preliminary results will be shared with the User and guidance provided to set or optimize the experimental conditions. This service will be performed by NF staff if the crosslinking reaction occurs at the home institution with guidance from us, or by a visiting User under our direct supervision if the entire workflow is requested.

The Structural Proteomics unit will perform crosslinking MS to characterize the topology of the protein-protein interactome of a particular target protein or enriched cellular fraction in situ. Examples include: interactome of pulldowns with an endogenously tagged protein of interest; interactome of vesicles/ cellular compartments; interactome of bacterial cells or virus/ host interactions after pulldown of specific virulence factors. Crosslinking reactions can be performed on cells, on lysates or after competitive or otherwise non-denaturing (native) elution. As these experiments require careful design, the User will be granted Access to preliminary acquisitions to characterise the sample and set the correct reaction conditions, or guidance to perform the optimization at their home institution. This service will be performed by NF staff in collaboration with a visiting User.

Structural biology and biochemical assays often require sample optimization to achieve stability and homogeneity, while understanding the oligomerisation state and precise composition of macromolecular complexes is crucial for unravelling their architecture. To address these needs, the Biophysics Unit offers a service leveraging various instruments to determine the molecular weights of species in solution and their thermal stability parameters. Techniques utilized include mass photometry, SEC-MALS, dynamic light scattering, and nanoDSF.

RNA sequencing is a powerful molecular biology technique used to analyse the transcriptome of a biological sample. The transcriptome refers to the complete set of RNA molecules, in particular messenger RNA (for mRNA sequencing) and/or non-coding RNAs (for total RNA sequencing), in a cell or tissue.

RNA sequencing is widely used in genomics research, functional genomics, and clinical studies to understand gene expression patterns, identify novel transcripts, and investigate how gene expression varies under different conditions. In addition, total RNA sequencing provides an insight also on the regulatory mechanisms underlying various biological processes.

This service enables researchers to explore the potential of single-molecule techniques and develop new protocols and workflows. Prior experience with dynamic single-molecule (DSM) approaches is not required.

The unit is equipped with two state-of-the-art instruments:

1. C-Trap ‘Dymo’ utilizes confocal scanning and is primarily used for imaging in solution.
2. C-Trap ‘Edge’ offers the flexibility to switch between wide-field fluorescence and TIRF imaging. Additionally, it allows label-free imaging via IRM, making it ideal for studies close to the surface.

The unit provides support for a wide range of experimental approaches.

This service will only be performed by NF staff, ideally accompanied by the User. As part of this service, the NF staff will check sample quality, troubleshoot, and define optimal conditions for data acquisition. If possible, an automation protocol for the experiment can be developed to streamline future data collection. The User can spend up to 10 days in the NF, testing various samples. The work can be split into two visits. The outcome of this service is an established protocol that can be used for ‘data acquisition’ service.

Microbiome analysis using 16S and ITS amplicon sequencing is a widely used technique to study the composition and diversity of microbial communities, particularly bacteria and fungi. The 16S ribosomal RNA (rRNA) gene is a molecular marker found in the genomes of bacteria and archaea, and its variable regions are commonly used for taxonomic classification, while ITS is used to profile fungal communities. Microbiome analysis using 16S and ITS amplicon sequencing is valuable in a range of fields, including environmental science, human health, agriculture, and more. It provides a cost-effective way to characterize microbial communities and understand their roles in various ecosystems or host-associated environments.

Abberior Facility Line STED with adaptive optics for deep-tissue super-resolution imaging.

• Available laser sources: 405 nm, 488nm, 561 nm and 640 nm
• Depletion laser: 775 nm
• Adaptive optics system
• Incubator (CO₂, temperature)
• MATRIX detector
• Available objectives:
• UPlanXApo 60x/1.42 Oil
• UPlanSApo 60x/1.20 Water