Labs that routinely do photolithography and soft lithography typically have what is called a plasma cleaner. A plasma cleaner has an electric coil, which creates a partially ionized gas consisting of electrons, ions, neutral atoms and molecules. The plasma creates reactive species on the surface of the glass and the device, which when placed together will form a permanent bond.
The plasma also makes the surface of the device hydrophilic, which facilitates the addition of liquids. In some cases, however, a permanent irreversible bond is not desirable. Also, many labs do not have access to a plasma cleaner.
To address this concern today will demonstrate reversible non plasma bonding of the neuron device to cover glass electrostatic forces between the PDMS and the glass forms a bond that is watertight and suitable for cell culture. I'd like to introduce you to Kiana Lee. She'll be doing the demonstrations Today.
Corning number 1 24 millimeter by 40 millimeter cover slips are cleaned using a water bath sonicate. The glasses are placed in a stainless steel rack that is then placed in a glass container filled with deionized water. The glass container is then placed into the water bath sonicate.
The glass slides are then sonicated for 30 minutes. After sonic deionized water is dumped out of the glass container. 70%ethanol is placed in the glass container with the glass slides, which is then brought inside a standard biosafety cabinet in order to keep them sterile.
After rinsing with 70%ethanol, the stainless steel rack is removed from the glass container and the glass slides allow it to dry completely for several hours or overnight. Next, while in the bowel safety cabinet, the glass slides are coated with PLL by soaking them in a container filled with PLL solution. The glass slides are allowed to soak in the PLL solution for a minimum of three hours or overnight.
After PLL coating, the PLL liquid is removed from the dish and then the glass slides are thoroughly rinsed with deionized water. After rinsing the PLL coated glass with water several times, water is then filled in the dish and the glass slides are allowed to soak in water for several hours. After soaking for several hours in deionized water, the water is aspirated from the dish.
The glass slides will be rinsed quickly and then the glass slides will be removed from the dish placed in a stainless steel rack and allowed to dry overnight In a biosafety cabinet, Neuron devices are brought into the biosafety cabinet and rinse thoroughly with 70%ethanol to sterilize them. The excess ethanol is then aspirated away from the device. The devices are then allowed to dry overnight in the biosafety cabinet the next day.
This sterile neuron device is simply placed on top of the clean, sterile and PLL treated glass. The PDMS forms a reversible bomb with the glass that is liquid tight. A device is bonded to the glass with gentle pressure cells, or media are then added immediately to the device.
Waiting too long to add media or cells to the device after the assembly can make it more difficult for the liquid to enter the device. It is important to note that the liquid does not readily enter the non plasma treated devices as fast as a plasma treated device. Plasma treated devices are initially hydrophilic just after plasma treated liquids readily flow into the hydrophilic device, whereas the flow through the non plasma bonded devices is much slower.
In some cases, it may be necessary to use a pipette to force liquid into the device. In this video, We have shown you how to non plasma bond the neuron device to cover glass. This is an efficient and inexpensive method to reversibly BLO, PDMS to glass surfaces.
We hope you have found this video informative and helpful. So that's it from the John Lab. Thanks for watching.
