Dear Jomar,

Did you check whether the functionalization of gelatin with methacryloyl groups was successful? 

In our latest work, we describe the complete process from GelMA synthesis and characterization, to application as cell culture models including various bio-assays on cell-laden hydrogels. The paper also contains a troubleshooting guide. Please find the link attached.

Regards,

Christoph

http://www.nature.com/nprot/journal/v11/n4/full/nprot.2016.037.html

A trouble-shooting question: When you UV cure the GelMA, are you sparging with inert gas? Dissolved oxygen can prevent the cross-linking reaction. This may be difficult because of the cells you want to keep alive, but necessary for the reaction to proceed. The GelMa may look to set because the GelMA can set temporarily just by being at a cold temperature due to gelatin physical crosslinks (Think like Jello desert) and these will break down with dilution and warming. Once chemically cross-linked, however, nothing will break the chemical bonds other than complete breakdown of the entire structure (only a strong oxidizer or something like that which cleaves the gelatin backbone would do this). I suspect something went wrong during your GelMA cure and you only have a physical cross-linked structure. Otherwise, DMEM shouldn't break it down. Hope this helps.

How long did you cure and what is the UV wavelength that you use? GelMA can be easily cure when it is a bit less viscous. Based on my experience, if 10% GelMA normally take longer time to cure compare to 5%. Moreover, the UV wavelength is really important depends on the absorb peak of your photo-initiator. I use irgacure with 5% gelma, it can cured within 120 seconds and it stayed in shape for a week (with cells). Normally, if you cure it properly, DMEM cannot wash it away.

I hope this help.

I cured it for 5 minutes, but I am not sure what the wavelength of the machine that we use is. I also use Irgacure 2959 with 10% GelMA.

There are times when I cured it for a longer period (10 mins) and the GelMA became white. Is this what we are trying to achieve?

Irgacure 2959 has absorption range 320-380 nm. If your machine does not provide in that range. GelMA will not be cure. For long curing time, the hydrogel might dry up and turn white. When GelMA is cured it still transparent or just translucent. 

irgacure 2959 info: http://www.xtgchem.cn/upload/20110629045632.PDF 

GelMA info please see

 3D Bioprinting of Vascularized, Heterogeneous Cell-Laden Tissue Constructs by David B. Kolesky et al.

http://www.xtgchem.cn/upload/20110629045632.PDF

Dear Jomar,

Did you check whether the functionalization of gelatin with methacryloyl groups was successful? 

In our latest work, we describe the complete process from GelMA synthesis and characterization, to application as cell culture models including various bio-assays on cell-laden hydrogels. The paper also contains a troubleshooting guide. Please find the link attached.

Regards,

Christoph

http://www.nature.com/nprot/journal/v11/n4/full/nprot.2016.037.html

Thank you all! Yes Christoph, I just noticed the new protocol by your team today, so I believe it will be very useful for troubleshooting GelMA production.

Thanks, Jomar!

Hi Jomar,

You want to check the wavelength and intensity of the light source you are using. You can check out a protocol here: https://biobots.io/biowiki/gelma/

Best,

Margaret