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Microinjection of Protein Samples Protocol - http://www.cshprotocols.org/cgi/content/abstract/2007/1/pdb.prot4657

Category: Microinjection Protocols: Microinjection of Proteins Protocols

This protocol describes a method for microinjecting proteins into the nucleus or cytoplasm of adherent cells. Microinjection equipment can be obtained from a number of suppliers; this protocol has been used with the Narishige IM-200 air pressure regulator and the Leitz micromanipulator. Using this system, it is possible to microinject a constant volume within a 50% difference among cells. - [Read Microinjection of Protein Samples Protocol]

Microinjection Setup Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/16/pdb.prot4396

Category: Microinjection Protocols: Microinjection Setup Protocols

This protocol describes the microinjection setup for injecting mouse zygotes to produce transgenic mice. - [Read Microinjection Setup Protocol]

Plating Cells for Microinjection Protocol - http://www.cshprotocols.org/cgi/content/abstract/2006/30/pdb.prot4650

Category: Cell Culture Protocols

Protocol describes a method for plating cells for microinjection onto etched coverslips. The coverslips for microinjection must be marked so that microinjected cells can be identified at time points after injection. - [Read Plating Cells for Microinjection Protocol]

Preparation (Pulling) of Needles for Gene Delivery by Microinjection Protocol - http://www.cshprotocols.org/cgi/content/abstract/2006/30/pdb.prot4651

Category: Microinjection Protocols: Microinjection Equipment

This protocol contains methods for pulling microinjection needles using two different models of pipette pullers. The advantage of pulling needles in the laboratory is that a variety of different needle types can be pulled, depending on the samples and cells being injected. An added advantage is cost; once a pipette puller has been purchased, boxes of glass capillaries are inexpensive compared to premade microinjection needles. - [Read Preparation (Pulling) of Needles for Gene Delivery by Microinjection Protocol]

Preparation and Loading of Protein Samples for Microinjection Protocol - http://www.cshprotocols.org/cgi/content/abstract/2007/1/pdb.prot4656

Category: Microinjection Protocols: Microinjection of Proteins Protocols

his protocol provides methods for the preparation of protein samples and for loading them into pulled microinjection pipettes. Stock solutions of proteins are thawed, diluted (if desired), centrifuged at high speed to remove aggregates, and kept on ice until loading. Loading into micropipettes can be done using either a "front-loading" or a "backfilling" procedure. - [Read Preparation and Loading of Protein Samples for Microinjection Protocol]

Preparation and Microinjection of x-Rhodamine-Labeled Vimentin Protocol - http://www.cshprotocols.org/cgi/content/abstract/2007/3/pdb.prot4676

Category: Cytoskeleton Protocols: Intermediate Filaments Protocols

Intermediate filaments (IF) are major cytoskeletal systems of vertebrate and many nonvertebrate cells whose expression is cell-type specific and developmentally regulated. This protocol describes the x-rhodamine labeling of one type of IF, vimentin, and a method for microinjection of the labeled vimentin into cultured cells. IF dynamics can then be examined with fluorescence microscopy. - [Read Preparation and Microinjection of x-Rhodamine-Labeled Vimentin Protocol]

Probing Protein Interactions Using GFP and FRET Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot3893

Category: Reporter Gene Assays: GFP Assay Protocols

In preparation for FLIM-FRET analysis, the appropriate donor and acceptor components must be introduced into live or fixed cells. The method of introduction depends on the nature of the components and the state of the cells. For example, plasmid DNAs encoding a protein of interest fused to a variant of GFP may be introduced into live cells by transfection or microinjection, whereas labeled antibodies are delivered by microinjection. - [Read Probing Protein Interactions Using GFP and FRET Protocol]

Purification of Bovine Lens and Bacterially Expressed Human Vimentin Protocol - http://www.cshprotocols.org/cgi/content/abstract/2007/3/pdb.prot4675

Category: Cytoskeleton Protocols: Intermediate Filaments Protocols

Intermediate filaments (IF) are major cytoskeletal systems of vertebrate and many nonvertebrate cells whose expression is cell-type specific and developmentally regulated. This protocol describes a method for purifying one type of IF, vimentin, from bovine lens tissue. Purification of human vimentin expressed in Escherichia coli is also described. These methods are useful in the preparation of other IF protein subunits for microinjection studies as well. - [Read Purification of Bovine Lens and Bacterially Expressed Human Vimentin Protocol]

Purification of Yeast Artificial Chromosome (YAC) DNA with Filtration Units Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/16/pdb.prot4393

Category: Yeast Protocols: Yeast Genetics Protocols: YACs Protocols

Protocol describes a method for purification of YAC DNA using ultrafiltration. This is the fastest method available, but care needs to be taken not to damage the DNA. The purified DNA can then be used for microinjection. - [Read Purification of Yeast Artificial Chromosome (YAC) DNA with Filtration Units Protocol]

Simple, Reliable Steps for DNA Fragment Isolation and Purification Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/16/pdb.prot4389

Category: Mouse Protocols: Mouse Genetics Protocols

Protocol describes a simple procedure for DNA fragment isolation and purification. The resulting DNA fragment can then. be used for microinjection to produce transgenic mice - [Read Simple, Reliable Steps for DNA Fragment Isolation and Purification Protocol]

Studying Mitosis in Cultured Mammalian Cells Protocol - http://www.cshprotocols.org/cgi/content/abstract/2007/3/pdb.prot4674

Category: Cell Biology and Cell Culture: Flow Cytometry Protocols: Cell Cycle Analysis Protocols

Protocol describes methods for maintaining healthy, dividing mammalian cells in culture and during imaging, when mitosis can be examined. Rose chambers are preferable for observation and microinjection of living mitotic cells, but slide/coverslip preparations are easy to make and do not require any special equipment. - [Read Studying Mitosis in Cultured Mammalian Cells Protocol]

Studying Mitosis in Cultured Mammalian Cells Protocol - http://www.cshprotocols.org/cgi/content/abstract/2007/3/pdb.prot4674

Category: Cytogenetics Protocols

Protocol describes methods for maintaining healthy, dividing mammalian cells in culture and during imaging, when mitosis can be examined. Rose chambers are preferable for observation and microinjection of living mitotic cells, but slide/coverslip preparations are easy to make and do not require any special equipment. Another inexpensive and easy-to-use alternative is to grow cells in a culture dish with a glass bottom. Such dishes are suitable for microinjection experiments. - [Read Studying Mitosis in Cultured Mammalian Cells Protocol]

Studying Mitosis in Cultured Mammalian Cells Prtocol - http://www.cshprotocols.org/cgi/content/abstract/2007/3/pdb.prot4674

Category: Cell Biology and Cell Culture: Cell Cycle Protocols: Mitosis Protocols

Protocol describes methods for maintaining healthy, dividing mammalian cells in culture and during imaging, when mitosis can be examined. Rose chambers are preferable for observation and microinjection of living mitotic cells, but slide/coverslip preparations are easy to make and do not require any special equipment. Another inexpensive and easy-to-use alternative is to grow cells in a culture dish with a glass bottom. Such dishes are suitable for microinjection experiments. - [Read Studying Mitosis in Cultured Mammalian Cells Prtocol]

Transgenic Outline: Mice and Rats - http://www.med.umich.edu/tamc/tgoutline.html

Category: Transgenic Mouse Protocols

Clone the Transgene, transgene Screening, Transgene Expression, DNA microinjection, Identify Transgenic Founders, Transgenic Analysis. Transgenic Animal Web. - [Read Transgenic Outline: Mice and Rats]

Xenopus Microinjection Protocol - http://tropicalis.berkeley.edu/home/manipulate_embryos/microinjection/inject.html

Category: Xenopus Protocols: Xenopus Injection Protocols

Protocol for Xenopus microinjection. - [Read Xenopus Microinjection Protocol]

Articles (1-4 of 4)

Tubulin Polymerization Protocol using GTP and GMPCPP

Tubulin is polymerized into microtubules by incubating tubulin at 37°C with GTP. A nucleation seed is added when the purpose is to assay microtubule elongation. Tubulin can also be polymerized for the purposes of recycling the tubulin or labeling the microtubules with fluorescently labeled tubulin. Based on the protocol by Timothy Mitchison of Harvard University.

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