SPI Supplies® Mica Sheets and Disks
Silanization of mica strips and glass and quartz slides
A number of protocols seem to exist for making substrates "sticky" in
terms of the adhesion of cells. The key "secret" ingredient is
3-aminopropyltriethoxysilane (3APTS).
- Dip the slides (or substrates) to be coated in a 2% 3APT in acetone
solution for one minute. The substrates must be as clean as possible,
which can be done by immersion in 20% hydrochloric acid in ethanol. An
alternative would be 20% aqueous sulfuric acid. The point is to make the
surface hydrophilic at this point. As further insurance that the surface
is cleaned to the degree necessary, they should then be immersed in
anhydrous acetone for two hours, the "anhydrous" being generated by storage
over molecular sieves for 24-36 hours.
- Prepare a 2% solution of 3APTS in anhydrous acetone (see above).
The substrates should be placed in an appropriate container(s) which could
be glass or polypropylene. The container should be filled completely with
the 2% solution and stored for 24 hours at 50°C or 4% solution for 12 hours.
The 3APTS silane treatment coats the substrates with amino groups.
- Dip the slides to be free of the 3APTS in 100% acetone for 1 minute
followed by:
- Dipping the slides in water for 1 minute.
- Repeat steps 3) and 4) but with agitation.
- Air dry in a dust free environment.
At this point the coated substrate should be "sticky" to the touch and ready
for use. They can be stored for about five days if immersed in distilled
water at 4.
Typical protocol for cell attachment:
The substrates are rinsed in the anhydrous acetone and immersed for one hour
in 1% EM grade glutaraldehyde for one hour. They are rinsed in acetone and
immersed for one hour in 1% glutaraldehyde. The carriers are then placed
into distilled water at 4°C. The surface should be now coated with
aldehyde groups, which would bond covalently with amino groups on the
surface of cells (or cell fractions). If cells (or cell fractions) must be
fixed before attachment (in glutaraldehyde), the 1% glutaraldehyde is
omitted. Exposed aldehyde groups on the fixed biological materials will
bond to the amino groups on the substrate.
The above information should be viewed as a composite of input we have
received from several researchers who do have differing views on exactly the
optimum way to conduct this kind of substrate preparation. We would
strongly suggest consulting the references given below.
Additional references:
- Angerer, L.M. and Angerer, R.C. Localization of mRNAs by in situ
hybridization. Functional Organization of the Nucleus: a Laboratory Guide.
Methods in Cell Biology, Vol 35, (Edited by B.A. Hamalko and S.C.R.
Elgin), pp. 37 - 71(1991).
- J. P. Robinson, P.Dunnill, and M. D. Lilly, Biochim. Biophys.
Acta 242, 659-661, (1971)
- M. Buechi and T. Baechi, J.Cell Biol. 83, 338-347, 1979.
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Tuesday December 02, 2008
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