User login
Credit: Virginia Tech
A novel technique that can detect the subcellular location of a protein may help improve the study of therapies for cancer and other diseases, according to a paper published in Chemical Science.
“Modulation of protein transport inside a cell is practiced as an important therapeutic approach for cancer treatment,” explained Chang Lu, PhD, of Virginia Tech in Blacksburg.
“The subcellular location of a target protein can also serve as a useful read-out for high-content screening of cancer drugs.”
With that in mind, Dr Lu and his colleagues set out to develop a simple and accessible protein detection method that can rapidly screen a large cell population and offers single-cell resolution.
Dr Lu noted that such techniques have been seriously lacking. For instance, fluorescence microscopy can only be used to analyze a limited number of cells.
And data collected by subcellular fractionation only reflects the average properties of the cell populations without revealing the heterogeneity that often exists among cells that seem identical.
Dr Lu and his colleagues had previously made some progress in screening cell populations using an electroporation-based technique, but it did not allow for the examination of native proteins and primary cells isolated from animals and from patients.
Their new work uses a method that “incorporates selective chemical release of cytosolic proteins with a standard procedure for fluorescent labeling of the protein,” Dr Lu said.
This simple tweak to the conventional cell-staining process allowed the researchers to pinpoint the subcellular location of the protein by measuring the amount of the residual protein after release. Using a flow cytometer, the speed of such measurement could reach 10,000 to 100,000 cells per second.
A key ingredient for the team’s process is saponin, an amphipathic glycoside. Saponin dissolves cholesterol and permeates the plasma membrane to allow protein release. And it “shows minimal effects on the state of the cell,” Dr Lu said. 
Credit: Virginia Tech
A novel technique that can detect the subcellular location of a protein may help improve the study of therapies for cancer and other diseases, according to a paper published in Chemical Science.
“Modulation of protein transport inside a cell is practiced as an important therapeutic approach for cancer treatment,” explained Chang Lu, PhD, of Virginia Tech in Blacksburg.
“The subcellular location of a target protein can also serve as a useful read-out for high-content screening of cancer drugs.”
With that in mind, Dr Lu and his colleagues set out to develop a simple and accessible protein detection method that can rapidly screen a large cell population and offers single-cell resolution.
Dr Lu noted that such techniques have been seriously lacking. For instance, fluorescence microscopy can only be used to analyze a limited number of cells.
And data collected by subcellular fractionation only reflects the average properties of the cell populations without revealing the heterogeneity that often exists among cells that seem identical.
Dr Lu and his colleagues had previously made some progress in screening cell populations using an electroporation-based technique, but it did not allow for the examination of native proteins and primary cells isolated from animals and from patients.
Their new work uses a method that “incorporates selective chemical release of cytosolic proteins with a standard procedure for fluorescent labeling of the protein,” Dr Lu said.
This simple tweak to the conventional cell-staining process allowed the researchers to pinpoint the subcellular location of the protein by measuring the amount of the residual protein after release. Using a flow cytometer, the speed of such measurement could reach 10,000 to 100,000 cells per second.
A key ingredient for the team’s process is saponin, an amphipathic glycoside. Saponin dissolves cholesterol and permeates the plasma membrane to allow protein release. And it “shows minimal effects on the state of the cell,” Dr Lu said.
Credit: Virginia Tech
A novel technique that can detect the subcellular location of a protein may help improve the study of therapies for cancer and other diseases, according to a paper published in Chemical Science.
“Modulation of protein transport inside a cell is practiced as an important therapeutic approach for cancer treatment,” explained Chang Lu, PhD, of Virginia Tech in Blacksburg.
“The subcellular location of a target protein can also serve as a useful read-out for high-content screening of cancer drugs.”
With that in mind, Dr Lu and his colleagues set out to develop a simple and accessible protein detection method that can rapidly screen a large cell population and offers single-cell resolution.
Dr Lu noted that such techniques have been seriously lacking. For instance, fluorescence microscopy can only be used to analyze a limited number of cells.
And data collected by subcellular fractionation only reflects the average properties of the cell populations without revealing the heterogeneity that often exists among cells that seem identical.
Dr Lu and his colleagues had previously made some progress in screening cell populations using an electroporation-based technique, but it did not allow for the examination of native proteins and primary cells isolated from animals and from patients.
Their new work uses a method that “incorporates selective chemical release of cytosolic proteins with a standard procedure for fluorescent labeling of the protein,” Dr Lu said.
This simple tweak to the conventional cell-staining process allowed the researchers to pinpoint the subcellular location of the protein by measuring the amount of the residual protein after release. Using a flow cytometer, the speed of such measurement could reach 10,000 to 100,000 cells per second.
A key ingredient for the team’s process is saponin, an amphipathic glycoside. Saponin dissolves cholesterol and permeates the plasma membrane to allow protein release. And it “shows minimal effects on the state of the cell,” Dr Lu said.