Graduation Year

2005

Document Type

Thesis

Degree

M.S.Ch.E.

Degree Granting Department

Chemical Engineering

Major Professor

Mark Jaroszeski, Ph.D.

Committee Member

Richard Gilbert, Ph.D.

Committee Member

Michael VanAuker, Ph.D.

Keywords

Corona, DC contact, Cell fusion, Melanoma cells, Fusion chamber

Abstract

From previous studies it has been demonstrated that the fusion of tumor cells with antigen-presenting cells generates hybrids that are known to induce anti-tumor immunity. With the advancement of scientific research and medicine, the need to produce cell-cell hybrids for cancer immunotherapy and for various other applications is substantial.

Among the many methods used to generate these hybrid cells, electrofusion is a technique that is more widely used and recognized as a method to efficiently produce hybrids. Electrofusion requires two steps. In the first step, cells are brought into close adjacent contact either by a mechanical method like centrifugation or by dieletrophoresis using alternating current (AC). The second step includes the reversible breakdown and fusion of cell membranes induced by high voltage direct current (DC) pulses.

The goal of this investigation was to study the use of electrical charge to bring cells into close contact with one another in the cell contact stage prior to delivering high voltage fusion pulses. The possibility of achieving considerable cell-cell contact was tested in two separate electrical systems.

In the first system B16 murine melanoma cancer cells were subjected to a range of direct current (DC) voltages between 4 V/cm and 40 V/cm. With the use of DC from a small power source the response of the cells was tested in multiple fusion chambers consisting of two or four electrodes. The configurations of the chambers were varied by changing the distance between the electrodes, the thickness, material and type of coating on the electrodes.

In the second system the movement of cells in the presence of corona charge was studied. B16 cells in a culture dish were confined by a circular grounded electrode and subjected to corona discharge for known periods of time. Application of corona charge (positive or negative) facilitated the contact of cells in the annular region between the two circular electrodes.

After series of tests, final designs for fusion chambers to be used with DC and with corona were developed. Cell contact achieved with the DC fusion chamber was not substantial enough to produce a significant amount of fusion yield. The fusion chamber designed to be used with corona on the other hand produced exceptional cell contact results consequentially generating fusion yields as high as 40%.

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