"We really have bridged the gap between different technologies, allowing us to do science on a whole new level", Chang Lu stated. In a study published in Analytical Chemistry, Chang Lu demonstrated that the technique can detect a handful of protein movements, or translocations, within entire populations of cells.
These movements are important to detect because they are involved in many disease processes, such as oncogenesis, wherein a normal cell becomes malignant, according to Robert Geahlen, a study co-author and researcher in the Department of Medicinal Chemistry and Molecular Pharmacology. "Protein translocations are involved in the activation of tumour cells", he stated. "Detecting these movements could help diagnose the type and stage of cancer in the future."
Chang Lu's method uses two existing technologies: electroporation - used to determine protein location - and flow cytometry, a technique capable of rapidly examining individual cells but blind to intracellular protein locations on its own.
The Purdue technique, called "electroporative flow cytometry", harnesses the discerning power of the first method with the speed of the second, according to Robert Geahlen. The method involves cells being sent through tiny channels within a microchip and undergoing electroporation, wherein electrical pulses open pores in cell membranes and protein is released from inside. Then, sensors measure protein concentrations. Since a protein's subcellular location can directly influence the amount of protein leaving the cell, as Chang Lu and Robert Geahlen have shown, this method is capable of indirectly determining location, as Robert Geahlen stated.
If proteins are in their original position, floating freely in the cell's interior, or cytoplasm, a large percentage of them will flow out of the cell upon electroporation, according to Chang Lu. If translocation has occurred, in which proteins migrate from the cytoplasm to tightly bind to the interior of the cell membrane, few will be able to leave.
Previous technologies detected either protein movement in a few individual cells via slow imaging techniques or took average measurements from larger groups of cells, data usually irrelevant to protein location in individual cells, as Chang Lu explained. "When looking at a few cells, you see the trees but not the forest", he stated. "When you take average measurements from large groups, you see the forest but not the trees. Our method allows us to know something about each tree in the forest."
The technology has the potential to be scaled up significantly, according to Chang Lu. In the study, 100-200 cells were processed per second, but that rate could potentially increase to speeds typical of flow cytometry, which goes through 10.000 cells per second. Speed increases can be achieved by optimizing the technology, Chang Lu stated.
The study examines the movement of a certain type of protein called kinases. Kinases and their translocations are important for activating and de-activating cells and sending signals to one another, according to Robert Geahlen. "There are many important kinases, enzymes and other proteins that become activated by translocation to the plasma membrane, and we've shown that we can detect one type of translocation", he stated. "It's a first step."
Chang Lu has filed a provisional patent for the technique and said that he could see it being used in a clinical setting in five to 10 years.
Study co-authors include graduate students Jun Wang, Leela Paris, Hsiang-Yu Wang, and postdoctoral researcher Ning Bao. Chang Lu and Robert Geahlen received funding from Purdue and the National Cancer Institute for the study. Chang Lu plans to further develop the technology in the future.