Preparing collagen and coating chip with collagen
We did a 1:1000 dilution of acetic acid (17.5 M) - we used 10 uL of acetic acid and 10 mL of sterile water.
We did a 2:3 dilution of collagen - we used 200 uL of 5 mg/mL collagen and 300 uL of the diluted acetic acid solution.
We coated Annikens chip (100 um) with collagen. We injected the collagen by using a 5 mL syringe and with a flow of around 50 uL/min. Stopped the flow afterwards and let the collagen settle for one hour. Aspirated the chip by using a 5 mL syringe multiple times. Then we used Elveflow pressure controller OB1 with nitrogen gas for about 30 minutes to dry the chip completely.
We casted two new chips - Annikens (100 um) and Thomas' (25 um).
We took out the chips from the oven after about 3 hours.
We passaged the cells as usual. For detail see here. We kept one flask and changed one flask. We took out 2 ml from the old flask into a 3 ml syringe.
Injecting cells into chip and preparing for timelaps
We used tubes marked "For Elveflow material" all day. We used hypodermic needle in size 0.60 all day (also for the collagen coating).
We filled the tube separately by using a 3 mL syringe manually with PBS and then used the same syringe to clean the chip with PBS (automatically). (This worked in terms of limiting bubbles inside the chip, but when we disconnected the syringe with PBS and the tube in order to switch to the syringe with cells, air came inside the tube and when we started injecting the cells there were a lot of big bubbles.)
We changed the syringe to a 3 mL syringe with cells in media. Kept the same tube. Injected the cells.
We changed the syringe to a 5 mL syringe with CO2 ind. media. Kept the same tube. Miscalculated and set the flow rate at 1 mL/min instead of 1 uL/min. Blew all the cells out.
We changed back to the 3 mL syringe with cells in media. Kept the same tube. A few cells came into the chip. They had probably attached to the inside of the syringe after having been inside the syringe for a while at that point. We found a small cluster of cells in one of the chambers.
We changed back to the 5 mL syringe with CO2 ind. media. Only 3 mL of media remained. This time we had included a Acrodisc filter between the syringe and the hypodermic needle to hopefully reduce the amount of air bubbles inside the chip.
We started a timelaps taking pictures of said cluster of cells every 1 hour for 3 days.