BOT/MICRO/ZOO 5364

SILANE

The silane compound comes as a liquid from Sigma. Its name is:3-aminopropyltriethoxysilane

The slides seem consistently sticky in our hands, although we once had trouble with really teeny weenie sections that someone here was cutting (but this was also for an application with lots of long rinses in running water). Further information about this can be found in:

Angerer, L.M. & Angerer, R.C. (1991) Localization of mRNAs by in situ hybridization. Functional Organization of the Nucleus: a Laboratory Guide. Methods in Cell Biology, Vol 35, (ed. By B.A. Hamalko & S.C.R. Elgin), pp. 37 - 71. Academic Press, San Diego.

--Tobias I. Baskin University of Missouri, Biological Sciences, Tucker Hall Columbia, MO 65211 USA voice: 314-882-0173

We would like to tell you about a method that we have now used routinely for about 5 years. It is based on procedures published by :

JP Robinson, P. Dunnill, and MD Lilly, in Biochim. Biophys. Acta 242, 659-661, 1971, and M. Buechi and T. Baechi, in J. Cell Biol. 83, 338-347, 1979.

  1. The glass (or other carriers) must be absolutely clean. This can be accomplished by immersing for several hours in aqueous 20% sulfuric acid, or 20% hydrochloric acid in ethanol. I prefer washing with Bon Ami cleaning powder until the surface is hydrophilic. The clean carriers are soaked for 2 h in anhydrous acetone that had been stored for at least 24 h over molecular sieve).
  2. Prepare a 2% solution of 3-aminopropyltriethoxysilane (Aldrich Chemical Co Milwaukee WI) in anhydrous acetone.
  3. The carriers are placed in appropriate containers (glass, Nalgene, polypropylene) which must be totally filled with the silane solution , and stored for 24 h at 50 degree C. If you use a 4% solution of silane, the carriers will be ready for use in 12 h. The silane treatment coats the carriers with amino groups.
  4. To attach fresh cells or cell fractions to the carriers, they are rinsed in acetone and immersed for one hour in 1% glutaraldehyde. The carriers are then placed into distilled water at 4 degree C. They can be kept for about 5 days. Their surface is now coated with aldehyde groups, which binds covalently with aminogroups on the surface of cells or cell fractions. If cells or cell fractions must be fixed before attachment (in glutaraldehyde fixative) the 1% glutaraldehyde is omitted. Exposed aldehyde groups on the fixed biological materials will bond to the amino groups on the carriers.

We have used sticky carriers prepared by this procedure to attach cells previously fixed in suspension to glass carriers:

--H. Ris and M. Malecki, Integrated Microscopy Resource, Univ. of Wisconsin, Madison WI. hris@facstaff.wisc.edu - Malecki@macc.wisc.edu