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Calibration of Micropipette Tips Protocols - http://www.cshprotocols.org/cgi/content/abstract/2007/1/pdb.prot4655

Category: Microinjection Protocols: Microinjection Equipment

This protocol describes an easy method for calibrating micropipette tips that have been pulled in the laboratory. It is essential to estimate the internal diameter of the pulled micropipette tip when adjusting parameters for a new puller or new type of glass tubing. A tip diameter of ~0.3 µm is optimal for the microinjection of mammalian cells in culture (e.g., CHO, PtK1, and COS-7). A 10% increase in diameter increases the delivery rate by more than 30% and can cause cell damage. - [Read Calibration of Micropipette Tips Protocols]

Cryogenic Preservation and Storage of Animal Cells Protocol - http://www.corning.com/Lifesciences/technical_information/techDocs/Cell_freezing_protocol.pdf

Category: Cell Culture Protocols: Cell Storing and Cell Freezing Protocols

Cryogenic preservation (storage below -100°C) of cell cultures is widely used to maintain backups or reserves of cells without the associated effort and expense of feeding and caring for them. The success of the freezing process depends on four critical areas: Proper handling and gentle harvesting of the cultures; Correct use of the cryoprotective agent; A controlled rate of freezing; Storage under proper cryogenic conditions. - [Read Cryogenic Preservation and Storage of Animal Cells Protocol]

Derivation of Trophoblast Stem (TS) Cell Lines from Blastocysts Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/18/pdb.prot4407

Category: Stem Cell Protocols

This protocol decribes derivation of TS cell lines from 3.5-days post coitum (dpc) mouse blastocysts. The procedure is similar to the derivation of embryonic stem (ES) cell lines. However, the success rate is considerably higher, and less expertise is required to recognize pluripotent TS cell colonies. - [Read Derivation of Trophoblast Stem (TS) Cell Lines from Blastocysts Protocol]

EMS Mutagenesis of Arabidopsis Seed Protocol - http://www.cshprotocols.org/cgi/content/abstract/2006/28/pdb.prot4621

Category: Plant Biology Protocols: Arabidopsis Genetics Protocols

EMS is used at concentrations that induce multiple point mutations in each plant, such that mutant alleles of a specific locus are found at a rate of ~1 in 2000-5000 M2 plants. This high rate of mutagenesis makes possible the screening of relatively few plants to find those with the phenotype of interest, a particular advantage if the screen is laborious or if only a small number of genes mutate to the required phenotype. - [Read EMS Mutagenesis of Arabidopsis Seed Protocol]

Erythrophagocytosis Assay - http://www.icp.ucl.ac.be/mexp/vipg_eph.html

Category: Immunology: Macrophage Monocyte Protocols

Erythrophagocytosis Assay. The erythrophagocytosis assay is performed to compare the phagocytic rate with and without anti-eythrocyte antibody in either macrophages from control animals or virus-infected counterparts. Andrei Musaji Viral Immunity and Pathogenesis Group - [Read Erythrophagocytosis Assay]

How to Measure and Monitor Linear Growth Rate - http://www.fgsc.net/neurosporaprotocols/How%20to%20measure%20and%20monitor%20line%20final.pdf

Category: Fungi Protocols: Neurospora Protocols

Information and tips on how to measure and monitor linear growth rate. - [Read How to Measure and Monitor Linear Growth Rate]

M5 Velocity (rate zonal) gradients for purification and assembly analysis of viruses. - http://www.axis-shield.com/densityhome/optiprep/M05.pdf

Category: Cell Biology and Cell Culture

Purification protocols of the viruses: HIV-1, Lassa virus, oncornavirus and other retroviruses. Protocol uses an iodixanol gradient in a sedimentation velocity mode to purifyHIV-1 virions without affecting the infectivity of the virus. In rate-zonal iodixanol gradients the HIV-1 was effectively separated both from Vif and from the microvesicles. - [Read M5 Velocity (rate zonal) gradients for purification and assembly analysis of viruses.]

MTT Cell Proliferation Assay Protocol - http://www.atcc.org/pdf/30-1010k.pdf

Category: Cell Biology and Cell Culture: Cell Viability Protocols

The MTT Cell Proliferation Assay measures the cell proliferation rate and conversely, when metabolic events lead to apoptosis or necrosis, the reduction in cell viability. The number of assay steps has been minimized as much as possible to expedite sample processing. The MTT Reagent yields low background absorbance values in the absence of cells. Includes: Determining optimal cell counts, performing an assay, data interpretations and troubleshooting. - [Read MTT Cell Proliferation Assay Protocol]

Native Aequorin - http://www.luxbiotech.com/Docs/Nanolight/Aequorin.pdf

Category: Signalling Signal Transduction Protocols: Calcium Measurement Protocols

Native Aequorin. NanoLight Technology. Aequorin has advantages over other Ca2+ indicators, for example, low leakage rate from cells, lack of intracellular compartmentalization or sequestration and it does not disrupt cell functions or embryo development. - [Read Native Aequorin]

Nucleic Acid Hybridization General Aspects - http://www.roche-applied-science.com/PROD_INF/MANUALS/InSitu/pdf/ISH_33-37.pdf

Category: Cytogenetics Protocols: In Situ Hybridization Protocols

Discusses the effects of various components of the hybridization solution on the rate of renaturation and thermal stability of DNA hybrids free in solution. Includes: The main parameters that influence hybridization; Additional hybridization variables; Competition in situ hybridization; Oligonucleotide hybridization; Standard in situ hybridization conditions. - [Read Nucleic Acid Hybridization General Aspects]

Ovary Cell Proliferation Test - http://embryo.ib.amwaw.edu.pl/invittox/prot/36.htm

Category: Pharmacology and Toxicology Protocols: Cytotoxicity Protocols: Cell Specific Cytotoxicity Assays Protocols

The rate of cellular proliferation may be regarded as an overall indicator of the physiological status of the cell. Therefore, the effect of various toxic substances on different cell functions will be reflected by changes in the proliferation rate. - [Read Ovary Cell Proliferation Test]

Passage of Embryonic Stem (ES) Cells Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/16/pdb.prot4401

Category: Stem Cell Protocols

This protocol describes passage of ES cells. They should be split at 1:3 to 1:7 every 2-3 days depending on their growth rate when they reach 70% confluency. They should never be allowed to grow past 90% confluency, but rather they should form tightly packed colonies not touching each other. - [Read Passage of Embryonic Stem (ES) Cells Protocol]

Selection of Appropriate Imaging Equipment and Methodology for Live Cell Imaging in Drosophila - http://www.cshprotocols.org/cgi/content/abstract/2006/29/pdb.ip21

Category: Microscopy Protocols: In Vivo Imaging Protocols

In the past two decades, there have been many revolutions in light microscopy techniques made possible by improvements in optics, detector technology, and computers. Furthermore, there is no indication that the rate of development of new equipment is slowing down. Here we attempt to provide an overview of available options and important considerations applicable to imaging Drosophila cells and tissues. - [Read Selection of Appropriate Imaging Equipment and Methodology for Live Cell Imaging in Drosophila]

Yeast Growth Rate Cytotoxicity Test - http://embryo.ib.amwaw.edu.pl/invittox/prot/33.htm

Category: Pharmacology and Toxicology Protocols: Cytotoxicity Protocols: Microorganism Cytotoxicity Assays Protocols

The proliferation rate of the yeast, Saccharomyces cerevisiae, may be regarded as an overall indicator of the physiological status of the cell. Therefore, the effect of various toxic substances on different cell functions will be reflected by changes in the rate of proliferation. It is possible to determine the toxicity of a test substance simply by measuring cell density. - [Read Yeast Growth Rate Cytotoxicity Test]

Articles (1-10 of 24)

Single Stranded Plasmid DNA Isolation Protocol

A Single Stranded Plasmid DNA Isolation Protocol describing the production and isolation of single-stranded DNA (ssDNA) using bacteriophagemid-containing bacteria and helper phage. Infection of the host cells with helper phage allows for packaging of ssDNA into bacteriophage. The ssDNA can then be isolated from phage particles.

Genomic DNA Labeling of Microarrays Protocol

DNA microarrays are an ordered arrangement of DNA molecules complementary to genes of interest that are "spotted" by robotic equipment onto a glass slide substrate. The expression of genes in cells can be monitored with microarrays by preparing cDNA from the mRNA of cells of interest and measuring the hybridization to the microarray. This protocol describes the labeling of genomic DNA for use as a probe for hybridization to the cDNA spotted on the array.

Microarray Protocol Preparation of Fluorescent DNA Probes from Human mRNA

This Microarray Protocol Preparation of Fluorescent DNA Probes from Human mRNA protocol describes the production of probes labeled with the fluorescent dyes, Cy3 and Cy5, following the synthesis of cDNA from human mRNA and the hybridization of the probes to DNA microarrays.

Probe Preparation for 22 x 40 mm DNA Microarrays

Probe Preparation for 22 x 40 mm DNA Microarrays Protocol.

Acid Guanidinium Thiocyanate RNA Isolation Phenol Chloroform Extraction

A single step RNA isolation protocol using Phenol Chloroform Extraction and Acid Guanidinium Thiocyanate. This RNA isolation method uses the fact that guanidinium thiocyanate can simultaneously lyse the cells and inactive cellular RNAses during the initial RNA isolation step allow a single step in the method.

Microinjection Pipettes Protocol

Microinjection Pipettes Preparation Protocol.

In Vitro Translated Xenopus Mos Kinase Assay Protocol

In Vitro Translated Xenopus Mos Kinase Assay Protocol. In response to progesterone, immature Xenopus oocytes mature to eggs that can be fertilized. The Mos protein kinase is essential for oocyte maturation, most likely due to its ability to activate the MAP kinase cascade. This MAP kinase cascade eventually leads to the activation of Cdc2/cyclin B and entry into M phase. In this protocol, tagged Mos kinase is translated in vitro, immunopurified, and used in a kinase assay.

Antibodies Phage Expression Protocol Using 96-well Microtiter Plates

A Phage display expression protocol for antibody expression in 96-well microtiter plates.

Immobilized Antigen Phage Antibody Selection Protocol

A protocol for the selection of Phage Antibodies using Immobilized Antigen. This method describes the selection of antibodies from bacteriophage antibody libraries that recognize a specific antigen. The phage display library of antibody-displaying phage particles is exposed to antigen attached to a solid substrate (Nunc Immuno™ tubes). The phage particles with affinity for antigen bind to the immobilized antigen and are selected from the library of phage expressing antibodies.