I am trying to make solid collagen gels to release from the plates as rafts. I am currently making them up as follows: 100ul MEM, 900ul diluted collagen, 20ul NaOH and 2ul bicarbonate.
Does anyone have any suggestions as to what i can do? If anyone has a protocol they know works i would be grateful if you could post it for me.
Thanks,
Lizz
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jeffreybarrett1
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Posted 13 February 2005 - 11:50 PM
Hi,
I have done a lot of work with making collagen gels. I would check your collagen concentration and possibly increase it. Also, check your end pH; if it is too low then you will not get proper gelling.
I use 8 parts collagen, 1 part reconstitution buffer, and 1 part 10x media. The reconstitution buffer consists of HEPES and sodium bicarbonate at the concentration in the 1x media plus 0.2N final conc. NaOH (in the buffer). Others have used down to 0.05N NaOH, but you have to see what works best for you. My final concentration of collagen is about 2.8 mg/ml, which seems to work well for growth of cells.
Please email me at j-barrett@northwestern.edu if you have further questions.
-Jeff
I have done a lot of work with making collagen gels. I would check your collagen concentration and possibly increase it. Also, check your end pH; if it is too low then you will not get proper gelling.
I use 8 parts collagen, 1 part reconstitution buffer, and 1 part 10x media. The reconstitution buffer consists of HEPES and sodium bicarbonate at the concentration in the 1x media plus 0.2N final conc. NaOH (in the buffer). Others have used down to 0.05N NaOH, but you have to see what works best for you. My final concentration of collagen is about 2.8 mg/ml, which seems to work well for growth of cells.
Please email me at j-barrett@northwestern.edu if you have further questions.
-Jeff
