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1.115 Aktuelle Fachpublikationen in CSH Protocols

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Preparation of Cellular Extracts from Xenopus Eggs and Embryos

01.06.2018 | Matthew C. Good; Rebecca Heald, CSH Protocols, 2018

Cell-free cytoplasmic extracts prepared from Xenopus eggs have been used extensively to recapitulate and characterize intracellular events in vitro. Egg extracts can be induced to transit the cell cycle and reconstitute assembly of dynamic structures including the interphase nucleus and the ...

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Isolation of IgY from Chicken Eggs

01.06.2018 | Fishman, J. B., Berg, E. A., CSH Protocols, 2018

Because IgY does not interact with either Protein A or Protein G, more traditional methods must be used for its isolation. To isolate chicken IgY from the yolk of chicken eggs, the yolks and the whites must be separated much in the way a cook does. Minimizing the egg white proteins is an ...

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In Vitro Fertilization in Mice

01.06.2018 | Takeo, T., Nakagata, N., CSH Protocols, 2018

The success of in vitro fertilization is strongly influenced by genetic background, handling of sperm and oocytes, and culture conditions. This protocol promotes enhanced rates of fertilization by using preincubation medium for sperm containing methyl-β-cyclodextrin (MBCD) and fertilization ...

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Analysis of Cell Viability by the alamarBlue Assay

01.06.2018 | Kumar, P., Nagarajan, A., Uchil, P. D., CSH Protocols, 2018

This protocol describes viability measurements for cell cultures in a 96-well tissue culture plate using alamarBlue (resazurin). The assay can be modified to accommodate larger plates; however, for a preliminary analysis of transfection reagents and parameters of the transfection protocol on cell ...

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Mouse Sperm Cryopreservation Using Cryoprotectant Containing L-Glutamine

01.06.2018 | Takeo, T., Nakagata, N., CSH Protocols, 2018

Efforts to advance sperm cryopreservation are ongoing and include modifications in the cryoprotective agents. The addition of l-glutamine maintains post-thaw motility and reduces plasma membrane damage to sperm.

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Ammonium Sulfate Fractionation of Antibodies

01.06.2018 | Fishman, J. B., Berg, E. A., CSH Protocols, 2018

Before the widespread availability and use of Protein A (rabbit) or Protein G (rodent) for the purification of IgG, the use of an ammonium sulfate "cut" was the standard method to isolate IgG and other serum proteins. The addition of ammonium sulfate reduces the effective solubility of proteins ...

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Isolation and Demembranation of Xenopus Sperm Nuclei

01.06.2018 | James W. Hazel; Jesse C. Gatlin, CSH Protocols, 2018

The inherent experimental advantages of intact amphibian eggs have been exploited for several decades to advance our understanding of fundamental developmental processes and the cell cycle. Characterization of these processes at the molecular level has been greatly advanced by the use of ...

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Analysis of Cell Viability by the Lactate Dehydrogenase Assay

01.06.2018 | Kumar, P., Nagarajan, A., Uchil, P. D., CSH Protocols, 2018

A common method for determining cytotoxicity is based on measuring the activity of cytoplasmic enzymes released by damaged cells. Lactate dehydrogenase (LDH) is a stable cytoplasmic enzyme that is found in all cells. LDH is rapidly released into the cell culture supernatant when the plasma ...

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Nucleus Assembly and Import in Xenopus laevis Egg Extract

01.06.2018 | Chen, P., Levy, D. L., CSH Protocols, 2018

Xenopus egg extract represents a powerful cell-free biochemical tool for studying organelle assembly and function. Large quantities of cytoplasm can be isolated, and biochemical manipulation of extract composition and cell cycle state is relatively straightforward. In this protocol, we describe ...

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Protein Quantitation

01.06.2018 | Greenfield, E. A., CSH Protocols, 2018

Methods for quantifying protein samples include colorimetric assays (e.g., using bicinchoninic acid, Bradford, or Lowry reagents) and ultraviolet (UV) detection (e.g., using NanoDrop technology). Seven such methods are outlined here.

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