I am trying Matrigel invasion assay for myeloma cell lines. I am using 8 microns membrane and coating the matrigel at 1: 3 conc. But am not getting results. I am using U266 cell line which is reported to have invasive potential when checked with IGF-1. But I am not getting any results in 48 hours. Some of the papers show pore size 10-12 um. I wonder if this could be a reason?
HI, what is your media composition and what is your concentration of matrigel (5mg/ml)? - we dilute it to 1 mg / ml. Also, we use a high FCS (up to 20%) as the chemoattractant in the lower chamber. I think 8 microns should be fine, it's what we have used in the past. What controls have you got, beads or another invasive line?
I would probably first test your chemoattractant concentrations in a chemotactic migration assay without matrix. I would also lower the Matrigel to a dilution of 1:20 or 1:50, coat for 1hr at 37C and then run the invasion for 24 or 48 hrs.
Thanks all for your suggestions. In fact I am using IGF(5ug/ml) and 20% FCS containing RPMI as chemoattractants. the composition of my matrigel is 1:3 as lower then that makes it very liquid kind of jelly. I am using U266 as a positive control and cells without chemoattractants in the lower chamber serve as treatment control. I will try lower conc of matrigel and will update if it works.
Thanks
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