western blot using culture media
Posted 09 March 2010 - 01:44 PM
i am currently trying to do western blots with culture media for proteins secreted into it. We use really small gels with a 3-4ul well capacity so i have been lyophilising the media samples then resuspending in 1.5ul and adding 1.5ul loading buffer. I have not seen any bands yet though and i know from mass spec that the protein i am probing for is there.
80ul of media takes ~2hrs to lyophilise and the vacufuge temp options are room temp, 30, 45, and 60 degrees. I use room temp, but after 2hrs the samples are hot to the touch. Is there a different method to lyophilise the protein? I don't know if the protein concentrating methods will reduce the volume enough to allow me to load the whole sample into 1 well.
Posted 09 March 2010 - 08:46 PM
Posted 09 March 2010 - 08:55 PM
Posted 16 March 2010 - 07:47 PM
Thanks for the help
Posted 17 March 2010 - 12:43 PM
genius does what it must
i do what i get paid to do