6/8/2014 Overview of Cell Lysis and Protein Extraction

http://www.piercenet.com/method/overview-cell-lysis-protein-extraction 1/3
Home > Protein Methods Library > Overview of Cell Lysis and Protein Extraction
Overview of Cell Lysis and Protein Extraction
Cell lysis is the first step in cell fractionation, organelle isolation and protein extraction
and purification. As such, cell lysis opens the door to a myriad of proteomics research
methods. Many techniques have been developed and used to obtain the best possible
yield and purity for different species of organisms, sample types (cells or tissue) and
target molecule or subcellular structure.
Page Contents:
Structure and Diversity of Cells
Cell Lysis and Protein Extraction
Cell Fractionation and Organelle Isolation
Protease Inhibition and Protein Stabilization
Protein Refolding
Structure and Diversity of Cells
All cells have a plasma membrane, a protein-lipid bilayer that forms a barrier separating
cell contents from the extracellular environment. Lipids comprising the plasma
membrane are amphipathic, having hydrophilic and hydrophobic moieties that associate
spontaneously to form a closed bimolecular sheet. Membrane proteins are embedded in
the lipid bilayer, held in place by one or more domains spanning the hydrophobic core.
In addition, peripheral proteins bind the inner or outer surface of the bilayer through
interactions with integral membrane proteins or with polar lipid head groups. The nature
of the lipid and protein content varies with cell type and species of organism.
Cell membrane structure.
Illustration of a lipid bilayer
comprising outer plasma membrane
of a cell.
In animal cells, the plasma membrane is the only barrier separating cell contents from
Related literature...
Cell Lysis Technical Handbook

Instructions | MSDS | CofA
Part No. Find
Share This Page

Share

Follow Us
Email Announcements
Follow
Like Share
Follow
Sign-in
Other Country
6/8/2014 Overview of Cell Lysis and Protein Extraction
http://www.piercenet.com/method/overview-cell-lysis-protein-extraction 2/3
the environment, but in plants and bacteria the plasma membrane is also surrounded by
a rigid cell wall. Bacterial cell walls are composed of peptidoglycan. Yeast cell walls are
composed of two layers of -glucan, the inner layer being insoluble to alkaline
conditions. Both of these are surrounded by an outer glycoprotein layer rich in the
carbohydrate mannan. Plant cell walls consist of multiple layers of cellulose. These
types of extracellular barrier confer shape and rigidity to the cells. Plant cell walls are
particularly strong, making them very difficult to disrupt mechanically or chemically.
Until recently, efficient lysis of yeast cells required mechanical disruption using glass
beads, whereas bacterial cell walls are the easiest to break compared to these other
cell types. The lack of an extracellular wall in animal cells makes them relatively easy
to lyse.
Cell Lysis and Protein Extraction
Historically, physical lysis was the method of choice for cell disruption and extraction of
cellular contents; however, it often requires expensive, cumbersome equipment and
involves protocols that can be difficult to repeat due to variability in the apparatus (such
as loose-fitting compared with tight-fitting homogenization pestles). Also, traditional
physical disruption methods are not conducive for high throughput and smaller volumes
typical of modern laboratory research.
In recent years, detergent-based lysis methods have become the norm. Through
empirical testing by trial and error, different detergent-based solutions composed of
particular types and concentrations of detergents, buffers, salts and reducing agents
have been developed to provide the best possible results for particular species and
types of cells. Detergents have both lysing and solubilizing effects.
Learn more...
Traditional Cell Lysis Methods
Detergents for Cell Lysis
Cell Lysis Solutions
Cell Fractionation and Organelle Isolation
By careful optimization of physical disruption techniques, detergent-buffer solutions and
density gradient methods, procedures have been developed to enable separation of
subcellular structures or classes of compounds. For example, with the appropriate
detergents, hydrophobic membrane proteins can be solubilized and separated from
hydrophilic proteins. A combination of tools and steps enable intact nuclei, mitochondria
and other organelles to be isolated for study or protein solubilization.
Learn more...
Cell Fractionation and
Organelle Enrichment
Protease Inhibition and Protein Stabilization
Cell lysis disturbs the carefully controlled cellular environment, allowing endogenous
proteases and phosphatases to become unregulated. As a result extracted proteins
become degraded or artifactually modified by the activities of these molecules.
To prevent these effects and obtain the best possible protein yield in cell lysis, protease
and phosphatase inhibitors are added to the lysis reagents. Numerous compounds have
Learn more...
Protease and Phosphatase
6/8/2014 Overview of Cell Lysis and Protein Extraction
http://www.piercenet.com/method/overview-cell-lysis-protein-extraction 3/3
been identified and used to inactivate or block the activities of proteases and
phosphatases by reversibly or irreversibly binding to them.
When the goal of cell lysis is to purify or test the function of a particular protein, special
attention must be given to the effects of the lysis reagents on the stability and function
of the protein(s) of interest. Certain detergents will inactivate the function of particular
enzymes, and long-term stability of extracted/purified proteins often requires that they
be removed from the initial lysis reagents and/or stabilized by addition of particular
compounds.
Inhibitors for Cell Lysis
View products...
Inhibitor Cocktails
Protein Stabilizing Cocktail
Protein Refolding
Some cell lysis and protein solubilization methods cause the denaturation of proteins.
Functional tests with such proteins requires that they be renatured. Many proteins
spontaneously refold into their native, functional structures when the denaturing
solubilization reagents are removed by dialysis. Other proteins, however, will fold into
non-functional and even insoluble forms by this process. In such cases, specialized
sets of buffer conditions must be tested to identify those that promote the highest
possible yield of properly refolded protein.
View products...
Protein Refolding Kit
Inclusion Body Solubilization
Reagent
Written and/or reviewed by Douglas Hayworth, Ph.D.