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- Erttmann, Saskia F., et al.
(författare)
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Protocol for isolation of microbiota-derived membrane vesicles from mouse blood and colon
- 2023
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Ingår i: STAR Protocols. - : CellPress. - 2666-1667. ; 4:1
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Tidskriftsartikel (refereegranskat)abstract
- Bacterial membrane vesicles have emerged as gadgets allowing remote communication between the microbiota and distal host organs. Here we describe a protocol for enriching vesicles from serum and colon that could widely be adapted for other tissues. We detail pre-clearing of serum or colon fluids using 0.2-μm syringe filters and their concentration by centrifugal filter devices. We also describe vesicle isolation with qEV size exclusion columns and finally the concentration of isolated vesicle fractions for downstream analyses. For complete details on the use and execution of this protocol, please refer to Erttmann et al. (2022).1
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- Giacomoni, Jessica, et al.
(författare)
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Protocol for optical clearing and imaging of fluorescently labeled ex vivo rat brain slices
- 2023
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Ingår i: STAR Protocols. - : Elsevier BV. - 2666-1667. ; 4:1
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Tidskriftsartikel (refereegranskat)abstract
- Tissue clearing is commonly used for whole-brain imaging but seldom used for brain slices. Here, we present a simple protocol to slice, immunostain, and clear sections of adult rat brains for subsequent high-resolution confocal imaging. The protocol does not require toxic reagents or specialized equipment. We also provide instructions for culturing of rat brain slices free floating on permeable culture inserts, maintained in regular CO2 incubators, and handled only at media change.
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- Saavedra Guillem, Beatriz, et al.
(författare)
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Selective quantitative N-functionalization of unprotected α-amino acids using NHC-Ir(III) catalyst
- 2023
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Ingår i: STAR Protocols. - : Elsevier BV. - 2666-1667. ; 4:2
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Tidskriftsartikel (refereegranskat)abstract
- Unnatural amino acids are valuable building blocks with numerous applications. Here, we present a quantitative technique for accessing mono-N-functionalized amino acids directly from unprotected substrates using alcohols as alkylating agents and an NHC-Ir(III) catalyst. We detail specific steps for catalyst preparation and application, as well as for catalyst recycling. The protocol excludes a few amino acids (l-cysteine, l-lysine, and l-arginine) and secondary alcohols.
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- Safi, Fatemeh, et al.
(författare)
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In vitro clonal multilineage differentiation of distinct murine hematopoietic progenitor populations
- 2023
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Ingår i: STAR Protocols. - : Elsevier BV. - 2666-1667. ; 4:1
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Tidskriftsartikel (refereegranskat)abstract
- Here we describe an in vitro co-culture system that can differentiate hematopoietic progenitor populations to all major hematopoietic lineages at clonal level. We present both a sensitive single-cell switch-culture system as well as a less laborious alternative barcoding protocol more convenient for larger cell numbers. Importantly, generation of all lineages from single long-term hematopoietic stem cells are described, following 21 days of culture. This protocol represents an efficient tool for validation experiments for single-cell genomics data. For complete details on the use and execution of this protocol, please refer to Safi et al. (2022).1
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- Wali, Gautam, et al.
(författare)
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Generation of human-induced pluripotent-stem-cell-derived cortical neurons for high-throughput imaging of neurite morphology and neuron maturation
- 2023
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Ingår i: STAR Protocols. - 2666-1667. ; 4:2
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Tidskriftsartikel (refereegranskat)abstract
- High-throughput imaging allows in vitro assessment of neuron morphology for screening populations under developmental, homeostatic, and/or disease conditions. Here, we present a protocol to differentiate cryopreserved human cortical neuronal progenitors into mature cortical neurons for high-throughput imaging analysis. We describe the use of a notch signaling inhibitor to generate homogeneous neuronal populations at densities amenable to individual neurite identification. We detail neurite morphology assessment via measuring multiple parameters including neurite length, branches, roots, segments and extremities, and neuron maturation.
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