Dear Smita Londhe,

AFM can be used at room temperature and atmospheric pressure and in general you don't need any special sample preparation for solid samples, but you should check the AFM system requirements. For convenience the sample should be flat at the bottom.

You should also have in attention the following:

- the AFM provides measurements of the surface heigh of your sample, for example, if you have features like spheres you can only 'see' the top part of the sphere;

- the area of measurement of the AFM is in general very small, like 100 x 100 microns or less and heigh is limited to few microns. You should check the AFM manual and if it is enough for your experiment.

- Assuming you have limited AFM time and you can not let the sample in the AFM for all the time it takes to grown the fungal, you need to find a way to mark the sample so that you can always return to the same position.

Good luck with your experiment.

AFM is a powerful tool for visualization and measuring of the mechanical properties in nano scale. Is it possible to scan a surface of native biological objects without any special sample preparation. Some AFM have a special chamber for visualization and force measuring in liquid.  

How big are your objects? The maximum size of scan area is nearly 200x200x5 microns (X x Y x Z), in a lot of cases it is less.

AFM is useful for visualization of a surface if the relief is not more than 1-2 microns. If it is higher, SEM and light microscopy are better.  

You can do next experiments for the growing fungi:

1. Visualization of a surface in nano scale for some stage of growth in tapping mode. You will capture a relief of fungi's surface. Probably, you will see some changes between the stages. For sample preparation it is enough to have just mechanical attached fungi on substrate. If your samples are too spiny it can be a problem for scanning.   

2. Measuring of the local mechanical characteristic of the fungi's surface in nano scale. By force spectroscopy technique you can capture some force-distance curves for interaction between an AFM probe's tip and a surface of fungi. It is possible to derive some parameters from these curves. Google "atomic force spectroscopy" and "peakforce AFM".

Good luck!      

Dear Smitha Londhe

Performing AFM on living cells is a challenging task. It requires to immobilize the sample, without denaturating the biological object. It is not so simple. A variety of solutions were discribed: electrostatic interactions, physical trapping, self assembling etc. You have to find the strategy that is adapted to your fungus.

for exemple you can read

Dague E, Alsteens D, Latgé JP, Dufrêne YF. High-resolution cell surface

dynamics of germinating Aspergillus fumigatus conidia. Biophys J. 2008 Jan

15;94(2):656-60. Epub 2007 Sep 21. PubMed PMID: 17890393; PubMed Central PMCID:

PMC2157220.

or

Formosa C, Pillet F, Schiavone M, Duval RE, Ressier L, Dague E. Generation of

living cell arrays for atomic force microscopy studies. Nat Protoc. 2015

Jan;10(1):199-204. doi: 10.1038/nprot.2015.004. Epub 2014 Dec 31. PubMed PMID:

25551664.

using this protocol we were able to follow C. albicans cells growing in the hyphal form (AFM experiments were conducted at 37°C, in MEM)

i recommand you this book

Advanced microscopy in Mycology (especially chapter 7 for AFM), Springer, T. Dahms and C. Czymmek Editors, 2015

Good luck with your experiments

Thanks you all.