post Protocols

Category: DNA Microarray Protocols

Post-Processing of Arrays. Brown Lab. - [Read Post-Processing of Arrays]

Category: Primary Cell Isolation and Culture Protocols: Mammalian Cell Culture Protocols

Cryopreserved PBMCs are a common specimen source for studies of immunological responses to vaccines, immunotherapies, etc. The health and viability of cells recovered post-cryopreservation are of course critical to the success and accuracy of immunological assays performed on them. This protocol standardizes PBMC isolation and cryopreservation techniques, specifically for the assessment of thawed cells by cytokine flow cytometry. - [Read Protocol for Isolation, Cryopreservation, and Thawing of PBMCs]

Category: Cytogenetics Protocols: In Situ Hybridization Tissue Sections Protocols

Protocol for RNA whole mount in situ hybridization. Includes: Embryo preparation; Prehybridization and Hybridization; Post-hybridization washes, blocking, and antibody incubation; Post-antibody washes; Color development. - [Read RNA Whole Mount In Situ Hybridization Protocol]

Category: Cell Biology and Cell Culture

Protocol for the isolation of the lipid-rich microdomains of the plasma membrane, notably caveolae and lipid rafts. Methods for the isolation of lipid rafts are based on the insolubility of these structures in the nonionic detergent TritonX-100. Either the intact cells are treated with a detergent-containing solution or a post-nuclear supernatant is prepared from a cell homogenate and then Triton X-100 is added to this supernatant. - [Read S20 Purification of detergent-insoluble lipid rafts from cells and tissues.]

Category: Cell Biology and Cell Culture: In Vitro Reconstitution Protocols

Information for protocol using single-tube, coupled transcription/translation reactions for eukaryotic in vitro translation. Includes information on: Translation Procedure; Positive Control Translation Reactions Using Luciferase; Cotranslational Processing Using Canine Pancreatic Microsomal Membranes; Post-Translational Analysis; Positive Control Luciferase Assays; Composition of Buffers and Solutions; Luciferase SP6/T7 Control DNAs - [Read Single-tube Coupled Transcription/Translation Reactions for Eukaryotic In Vitro]

Category: RNAi Technology Protocols

Protocol Involves the transfection of siRNA into RAW 264. 7 cells using Lipofectamine 2000.  Cells are transfected with siRNA twice (on subsequent days).  Target gene knockdown is assessed from total RNA isolated 48 hr post-transfection or from protein isolated 72 hr post-transfection. - [Read siRNA Double Transfection of RAW 264.7 Cells with Lipofectamine Protocol]

Category: RNAi Technology Protocols

Protocol Involves the transfection of siRNA into RAW 264. 7 cells using Lipofectamine 2000.  Cells are transfected with siRNA twice (on subsequent days).  Target gene knockdown is assessed from total RNA isolated 48 hr post-transfection or from protein isolated 72 hr post-transfection. - [Read siRNA Double Transfection of RAW 264.7 Cells with Lipofectamine Protocol II]

Category: Lipid Protocols

Protocol for sphingomyelin quantitation post-choline labeling of HL60 cells. - [Read Sphingomyelin Quantitation Post-choline Labeling of HL60 Cells Protocol]

Category: Primary Cell Isolation and Culture Protocols

This protocol describes a method for static culture of early postimplantation mouse embryos on a microscope stage. Embryos between 6.5 and 9.5 days post coitum (dpc) can be cultured and imaged for 24 hours, with very little growth retardation. - [Read Static Culture of Postimplantation Embryos for Imaging Protocol]

Category: Mouse Protocols: Mouse Cell Culture Protocols

Protocol describes a method for static culture of early postimplantation mouse embryos on a microscope stage. Embryos between 6.5 and 9.5 days post coitum (dpc) can be cultured and imaged for 24 hours, with very little growth retardation. - [Read Static Culture of Postimplantation Embryos for Imaging Protocol]

Category: Cell Culture Protocols

Protocol describes static culture of postimplantation embryos, an alternative to the roller method. The static method is best suited to 6.0 to 7.0 days post coitum (dpc) embryos followed for 24 hours (7.0 dpc embryos) to 48 hours (6.0 dpc embryos) of development. It allows repetitive real-time observation with minimal handling of the embryo. It is especially useful if single or small groups of embryos need to be distinguished from each other. - [Read Static Culture of Postimplantation Embryos Protocol]

Category: DNA Protocols: DNA Sequencing Protocols: LICOR Sequencing Protocols

Protocol for tracking and post- sequence processing data from LICOR. - [Read Tracking and Post-Sequence Processing Data from Licor Protocol]

Category: Mouse Protocols: Mouse Reproductive System Surgery Protocols

Protocol describes a procedure for uterine transfer, which is used for chimera production. The method is based on extensive work which resulted in the first successful development and birth of in-vitro-cultured mouse embryos. It is best to practice this procedure first on a cadaver and then on an anesthetized 2.5-days post coitum (dpc) pseudopregnant mouse using blue Affigel beads rather than embryos. - [Read Uterine Transfer Protocol]

Category: Cytogenetics Protocols: In Situ Hybridization Tissue Sections Protocols

Protocol for visualizing miRNAs in adult skeletal muscle by hybridization in situ. Includes: Visualizing miRNAs in adult skeletal muscle by hybridization in situ; Probe preparation; Tissue preparation; Fixation; Deproteinization with Proteinase K; Acetylation of the tissue; Dehydration; Hybridization step; Post-Hybridization Washes (Step 1-6 should be performed on shaker); Signal Detection (Steps should be performed on a shaker); Post-Detection Washes. - [Read Visualizing miRNAs in Adult Skeletal Muscle by Hybridization In Situ Protocol]

Category: Cytogenetics Protocols: RNA In Situ Hybridization Protocols

Protocol for visualizing miRNAs in adult skeletal muscle by hybridization in situ. Includes: Visualizing miRNAs in adult skeletal muscle by hybridization in situ; Probe preparation; Tissue preparation; Fixation; Deproteinization with Proteinase K; Acetylation of the tissue; Dehydration; Hybridization step; Post-Hybridization Washes (Step 1-6 should be performed on shaker); Signal Detection (Steps should be performed on a shaker); Post-Detection Washes. - [Read Visualizing miRNAs in Adult Skeletal Muscle by Hybridization in situ Protocol]

Category: C. elegans Protocols

Protocol for whole-mount in situ hybridization for the detection of RNA in C. elegans embryos. Includes: Collection of Embryos; Permeabilization and fixation of embryos; Prehybridization; Hybridization; Post-hybridization washes; Probe Detection; Double-labeling; Interpretation. - [Read Whole-Mount In Situ Hybridization for the Detection of RNA in C. elegans Embryos Protocol]