Hi there,

Values will differ based on the state of gel hydration and what it's being tested in.

I'll ask some relevant questions (rhetorically) that might help.

Was it swollen/highly-hydrated? Was the gel compressed in any way or was there loss of hydration during the test (or did it self-collapse in any manner)?  Was the collagen pepsin-digested or acid-solubalized? What solvent(s) was(were) used? Was the test conducted in an aqueous environment (i.e. submerged or replenished with a form of medium or PBS)? What was the osmolarity of solutes or viscosity in/of that medium? Are there pictures in the publication that indicate proper necking? Did the gels look like they were self-collapsing on one of the grips (unevenly)?

I hope that helps.

Regards,

Mark

Hi Mark,

The collagen gel was prepared as follows:

Collagen solution dissolved in acetic acid was neutralized by adding PBS, NaOH and distilled water. The solution was gelled at 37 deg for 40 min on AFM substrate. The gel was intact and not very swollen. It was maintained in the hydrated environment in PBS and AFM stiffness measurement was done. Loss of water from the gel by evaporation was minimal.

What would be your thought on this? 

Thank you.

Sunil

Hi,

Are you sure that comparing the results of your method with reported results on compression testing is meaningful? I mean, the scale of your measurements is different from what is reported in the literature and collagen hydrogels have multiscale properties if you consider the stiff fibers forming a porous hydrated network, so you might need to perform some statistical work to validate your AFM data with the results of compression testing.

The processing conditions seriously affect the mechanical properties of the formed hydrogel, so you should minimize the differences between your method and your references and if there are inevitable differences, at least  their particular effects should be considered. 

Col source will also affect the stiffness because of the structural differences (you can see that by comparing the intensity of amide 1&2 groups in FT-IR), even though you have the same Col type. So you should also use the same source, as you might already know :)

I have seen a few papers reporting Col mechanical properties by AFM, which makes your work valuable. I found the following article interesting and it might help you in your work.

Best,

Ali

Article Multi-scale Mechanical Characterization of Highly Swollen Ph...

 Thanks Ali for your descriptive suggestions. I really appreciate it. 

The paper you posted is bovine collagen type1. I wonder if there is any paper on determining the stiffness of collagen type1 rat tail gel using AFM because I have found none. 

Thanks again.

Sunil

You may want check the following discussions: https://www.researchgate.net/post/How_to_make_my_rat_tail_collagen_I_gel https://www.researchgate.net/post/Can_anyone_suggest_me_the_buffer_conditions_or_methods_for_solubilising_collagen_powder_other_than_acidic_condition

Hi again,

Sorry for my delayed response.

During my time with collagen, I recall there being a lot of difficulty distinguishing if the AFM tip was generating readings from media or fibres. With one such setup, distinguishing between inter-fibrillar voids and actual collagen, was indeed a unique task inspiring many thoughts.

I do remember being taught to question the robustness of every published result. If you are showing something with certainty, that goes against precedents set in the literature, definitely reach out to those academics whose names are cited on previous work. Try to get them to articulate the means by which their results were obtained. This might help navigate you in further steps/decisions you may wish to pursue. Don't dismiss the validity of your data quite yet; continue to keep reaching out to people to see what you will get.

Regards,

Mark James-Bhasin

Thanks Mark.

Hey Sunil,

We just published a paper characterizing Stiffness of collagen (type 1 rat tail) gel (concentration 1-5 mg/ml) using AFM.

Check this link, Article Three‐dimensional collagenous niche and azacytidine selectiv...

In supplementary section I explained the experimental set up as well.