Trying to get force curves on collagen, but having trouble

lmoore38 posted on Mon, Jul 11 2011 10:53 AM

Hello, I am a graduate student at Saint Louis University working with the AFM.  I am trying to get force curves on a collagen hydrogel over a period of 5 days, while a capillary network develops.  Right now, I am just trying to get a force curve on my collagen sample with no cells.  We have a Novascan machine, and I am using spherical tips with a spring constant of 0.6 N/m. I am running into a few problems that I was hoping I might find an answer to here.

1.  We were finding that when trying to position the probe over the sample, the very back of the cantilever was submerging into the collagen, thus not allowing the actual tip to function properly.  To solve this, I used less collagen on the coverslip, and tried to position the probe so that just the very tip was over the collagen itself.  We did get a force curve that looked similar to those in literature, but I'm not sure if it is actually reading the glass underneath the collagen.  We are so close to the edge, that I'm wondering if I'm reading collagen or glass beneath it since the sample is so thin at the edge. 

2.  When we tried to position the probe just a little further in to read a thicker part of the gel, we are finding that the jump-to-contact forces are so strong, that even if the back of the probe doesn't submerge, the tip itself is sucked into the gel and can't come out.  We've tried withdrawing a few microns each time, but the same thing keeps happening. 

Is my sample too soft?  Do you think these tips are ok?  Do I need to fix the sample if I want any good readings?  I was thinking that submerging the sample in PBS or something would help eliminate those jump-to-contact forces that seem to be messing up my data, but wanted to get a few professional opinions on if this is a good idea or not. 

Thank you so much for your time!

Lisa